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  • 501.
    Huijbers, Elisabeth J. M
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Development of a Cancer Vaccine Targeting Tumor Blood Vessels2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    A treatment strategy for cancer is the suppression of tumor growth by directing an immune response to the tumor vessels, which will destroy the tissue.

    In this thesis we describe the development of a vaccine that targets antigens expressed around angiogenic vasculature in most solid tumors. These antigens are alternative spliced extra domains of glycoproteins present in the extracellular matrix; e.g. the extra domain-B (ED-B) and extra domain-A (ED-A) of fibronectin and the C-domain of tenascin-C (TNCC).

    We show that it is possible to break self-tolerance and induce a strong antibody response against ED-B by vaccination. Furthermore, tumor growth was inhibited and the changes observed in the tumor tissue were consistent with an attack of the tumor vasculature by the immune system.

    For clinical development of therapeutic vaccines, targeting self-molecules like ED-B, a potent but non-toxic biodegradable adjuvant is required. The squalene-based Montanide ISA 720 (M720) in combination with CpG DNA fulfilled these requirements and induced an equally strong anti-self immune response as the preclinical golden standard Freund’s adjuvant. We have further characterized the immune response against ED-B generated with the adjuvant M720/GpG. 

    The ED-B vaccine also inhibited tumor growth in a therapeutic setting in a transgenic mouse model of pancreatic insulinoma in which tumorigenesis was already initiated. Furthermore, antibodies against ED-A and TNCC could be induced in mice and rabbits. We analyzed the expression of ED-A in breast tumors of transgenic MMTV-PyMT mice, a metastatic breast cancer model, with the aim to use this model to study the effect of an ED-A vaccine on metastasis. We also detected ED-B in canine mammary tumor tissue. Therefore vascular antigens might also represent potential therapeutic targets in dogs. 

    All together our preclinical data demonstrate that a vaccine targeting tumor blood vessels is a promising new approach for cancer treatment. 

  • 502.
    Huijbers, Elisabeth JM
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Femel, Julia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Cedervall, Jessica
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hein, Tobias
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Hellman, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Olsson, Anna-Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Development of a therapeutic vaccine targeting blood vessels in primary tumors and metastases2012Manuscript (preprint) (Other academic)
  • 503. Hulspas, Ruud
    et al.
    Villa-Komaroff, Lydia
    Koksal, Erin
    Etienne, Kenol
    Rogers, Patricia
    Tuttle, Matt
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Sharpe, John
    Berglund, David
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Transplantation Surgery.
    Purification of regulatory T cells with the use of a fully enclosed high-speed microfluidic system2014In: Cytotherapy, ISSN 1465-3249, E-ISSN 1477-2566, Vol. 16, no 10, p. 1384-1389Article in journal (Refereed)
    Abstract [en]

    Background aims. Despite promising advances in cellular therapies, it will be difficult to fully test or implement new therapies until advances are made in the processes for cell preparation. This study describes the use of an advanced prototype of a flow-cytometry cell purification system constructed for operation in a clinical environment to prepare regulatory T cells defined as CD4(+)/CD25(bright)/CD127(neg/low). Methods. The sort performance of the Gigasort system was directly compared with available droplet sorters using mixtures of highly fluorescent and non-fluorescent 5-mu m polystyrene particles. CD4(+)-enriched cell preparations were processed with the use of a sterile, disposable fluid handling unit with a chip containing parallel microfluidic-based sorters. Results. Similar purity and sort efficiency as found with droplet sorters were obtained with the 24-channel chip sorter system. Starting with 450 million fresh peripheral blood mononuclear cells, 150,000 to 1.7 million cells that were, on average, 85% FoxP3-positive and 97% viable, were obtained in <4 h. Conclusions. This study presents a technology adapted to regulatory requirements for clinical cell purification and that achieves high throughput and cell-friendly conditions by use of a microfluidic chip with 24 parallel microsorters, providing a rapid, sterile method of purifying regulatory T cells accurately and with excellent viability.

  • 504.
    Hultgren Hörnquist, Elisabet
    Örebro University, School of Medicine, Örebro University, Sweden.
    The mucosal immune system in microscopic colitis2012In: Microscopic colitis / [ed] S. Miehlke, A. Münich, Basel: S. Karger, 2012, p. 33-39Conference paper (Other academic)
  • 505.
    Hylén, Ulrika
    et al.
    Örebro University, School of Medical Sciences. University Health Care Research Center.
    Eklund, Daniel
    Örebro University, School of Medical Sciences.
    Humble, Mats B.
    Örebro University, School of Medical Sciences. University Health Care Research Center.
    Bartoszek, Jakub
    School of Medical Sciences, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Särndahl, Eva
    Örebro University, School of Medical Sciences.
    Bejerot, Susanne
    Örebro University, School of Medical Sciences. University Health Care Research Center.
    Increased inflammasome activity in markedly ill psychiatric patients: An explorative study2020In: Journal of Neuroimmunology, ISSN 0165-5728, E-ISSN 1872-8421, Vol. 339, article id 577119Article in journal (Refereed)
    Abstract [en]

    The aim of this study was to investigate inflammatory perturbations in 40 patients with severe and complex psychiatric disorders by studying the activity of the NLRP3 inflammasome, with a trans-diagnostic approach. Gene expression of CASP1, NLRP3, PYCARD, IL1B, IL1RN, TNF showed a significant increase in the patient group compared to a matched control group. Plasma levels of IL1Ra, IL-18, TNF, IL-6 and CRP were increased in the patient group. Within the patient group, increased gene expression of inflammatory markers correlated with increased disease severity. The findings support the inflammation hypothesis for markedly ill psychiatric patients across diagnostic groups.

  • 506.
    Hägglund, Hans
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Remberger, M.
    Ringden, O.
    Twenty-year follow-up of a randomized trial comparing intraosseous and i.v. BM transplantation2014In: Bone Marrow Transplantation, ISSN 0268-3369, E-ISSN 1476-5365, Vol. 49, no 12, p. 1541-1542Article in journal (Refereed)
  • 507.
    Häggqvist, Susana
    Linköping University, The Department of Physics, Chemistry and Biology.
    Effects of different conditions of HIV-1 on plasmacytoid dendritic cells in maturation and function2008Independent thesis Basic level (degree of Bachelor), 10 points / 15 hpStudent thesis
    Abstract [en]

    Plasmacytoid dendritic cells (PDCs) are one cellular target of HIV-1 and respond to the virus by producing type I interferons and chemokines. PDCs exposed to HIV-1 strongly upregulate the expression of maturation markers such as CD83, CD80, CD86 and CCR7, which will turn them into professional antigen presenting cells with the ability to stimulate naïve CD4+T cells. When HIV-1 binds to the CD4 receptor and a co-receptor (CCR5 or CXCR4) on PDCs, the cell takes up the virus by endocytosis. In response to this, PDCs will become activated and express maturation markers on their surface that make them able to stimulate T cells to trigger an immune response. In this thesis, studies have been performed with different forms of HIV-1, i.e. opsonized virions covered in complement and antibodies since these forms are supposed to be more similar to how HIV appears in the body. According to our results there is no significant difference in PDC maturation between the free and opsonized HIV-1.

  • 508.
    Hässler, Signe
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Sciences. CKMF.
    Ramsey, Chris
    Marits, Per
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Sciences. CKMF.
    Kämpe, Olle
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Sciences. CKMF.
    Surh, Charles D.
    Peltonen, Leena
    Winqvist, Ola
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Sciences. CKMF.
    Increased antigen presenting cell-mediated T cell activation in mice and patients without the autoimmune regulator2006In: European Journal of Immunology, Vol. 36, no 2, p. 305-317Article in journal (Refereed)
    Abstract [en]

    Patients with autoimmune polyendocrine syndrome type I (APS I)suffer from endocrine and non-endocrine disorders due to mutations in the autoimmune regulator gene (AIRE). Mouse Aire is expressed both in thymic medullary epithelial cells and in peripheral antigen-presenting cells, suggesting a role in both central and peripheral tolerance. We here report that Aire(-/-) dendritic cells (DC) activate naive T cells more efficiently than do Aire(+/+) DC. Expression array analyses of Aire(-/-) DC revealed differential regulation of 68 transcripts, among which, the vascular cell adhesion molecule-1 (VCAM-1) transcript was up-regulated in Aire(-/-) DC. Concurrently, the expression of the VCAM-1 protein was up-regulated on both Aire(-/-) DC and monocytes from APS I patients. Blocking the interaction of VCAM-1 prevented enhanced Aire(-/-) DC stimulation of T cell hybridomas. We determined an increased number of DC in spleen and lymph nodes and of monocytes in the blood from Aire(-/-) mice, and an increased number of blood monocytes in APS I patients. Our findings imply a role for Aire in peripheral DC regulation of T cell activation, and suggest that Aire participates in peripheral tolerance.

  • 509.
    Hårdstedt, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Studies of Innate and Adaptive Immunity in Islet Transplantation2014Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Clinical islet transplantation is today an established alternative treatment for a selected group of type 1 diabetes patients. The predominant technique for transplantation is infusion of islets in the liver via the portal vein. Obstacles to advancing islet transplantation include limited engraftment resulting from an immediate blood-mediated inflammatory reaction (IBMIR), a life-long need for immunosuppression and the shortage of organs available.

    In this thesis, innate and adaptive immunity were explored in allogeneic and xenogeneic settings, with the long-term goal of preventing islet graft destruction. Methods for studying immune responses to islets in blood and engrafted islets in liver tissue (intragraft gene expression) were developed and refined. The innate response to human islets and exocrine tissue in ABO-compatible blood was characterized up to 48 h using a novel whole-blood model. Physiological changes in the blood during incubations were explored and adjusted to allow prolonged experiments. Increased production of chemokines targeting CXCR1/2, CCR2 and CXCR3 was observed, accompanied by massive intra-islet neutrophil infiltration. Notably, endocrine and exocrine tissue triggered a similarly strong innate immune response.

    Two studies of adult porcine islet transplantation to non-human primates (NHPs) were performed. Expression of immune response genes induced in liver tissue of non-immunosuppressed NHPs (≤72 h) was evaluated after porcine islet transplantation. Up-regulation of CXCR3 mRNA, together with IP-10, Mig, MIP-1α, RANTES, MCP-1 and cytotoxic effector molecule transcripts, was associated with T-cell and macrophage infiltration at 48-72 h. Long-term survival (>100 days) of adult porcine islets in a NHP model was later demonstrated using T-cell-based immunosuppression, including co-stimulatory blockade (anti-CD154 mAb). Graft failure was associated with increased levels of circulating, indirectly activated T cells, non-Gal pig-specific IgG and gene transcripts of inflammatory cytokines. Microarray analysis of the response to inflammatory cytokines in cultured porcine islets identified genes involved in cell death, immune responses and oxidative stress; this gene pattern coincided with physiological changes (decrease in insulin and ATP content).

    In summary, allogeneic whole-blood experiments and xenogeneic in vivo studies underscored the importance of preventing early inflammation and cell-recruitment to avoid islet graft loss in islet transplantation. Long-term survival of porcine islets in NHPs was shown to be feasible using T-cell-directed immunosuppression, including anti-CD154 mAb.

  • 510.
    Hårdstedt, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Finnegan, Colleen P
    Kirchhof, Nicole
    Hyland, Kendra A
    Wijkstrom, Martin
    Murtaugh, Michael P
    Hering, Bernhard J
    Post-transplant upregulation of chemokine messenger RNA in non-human primate recipients of intraportal pig islet xenografts2005In: Xenotransplantation, ISSN 0908-665X, E-ISSN 1399-3089, Vol. 12, no 4, p. 293-302Article in journal (Refereed)
    Abstract [en]

    BACKGROUND:

    We have previously shown that pig-to-primate intraportal islet xenografts reverse diabetes, escape hyperacute rejection, and undergo acute cellular rejection in non-immunosuppressed recipients. To gain a better understanding of mechanisms contributing to xenoislet rejection in non-human primates we examined gene expression in livers bearing islet xenografts in the first 72 h after transplantation.

    METHODS:

    Liver specimens were collected at sacrifice from seven non-immunosuppressed rhesus macaques at 12, 24, 48 and 72 h after intraportal porcine islet transplantation. Following total RNA extraction, mRNA was quantified using SYBR green real-time reverse transcription polymerase chain reaction (RT-PCR) for species-specific immune response genes. Data were analyzed using comparative cycle threshold (Ct) analysis, adjusted for specific primer-efficiencies and normalized to cyclophilin expression.

    RESULTS:

    Porcine insulin mRNA was detected in all liver samples. Cluster analysis revealed differential gene expression patterns at 12 and 24 h (early) compared with at 48 and 72 h (late) post-transplant. Gene expression patterns were associated with histological findings of predominantly neutrophils and only a few lymphocytes at 12 and 24 h and an increasing number of lymphocytes and macrophages at 48 and 72 h. Transcript levels of CXCR3 and its ligands, interferon-inducible protein 10 (IP-10) and monokine induced by IFN-gamma (Mig), significantly increased between early and late time points together with expression of MIP-1alpha, regulated on activation normal T expressed and secreted protein (RANTES) and MCP-1. CCR5 showed only a marginal, non-significant increase. Fas ligand, perforin and granzyme B transcripts were all elevated at 48 and 72 h post-transplant.

    CONCLUSIONS:

    Our data suggest that CXCR3, with ligands IP-10 and Mig, is involved in T cell recruitment in acute islet xenograft rejection in non-human primates. Upregulation of RANTES and MIP-1alpha transcripts in the absence of a significant CCR5 increase suggests a possible involvement of other chemokine receptors. MCP-1 expression is associated with T cell and macrophage infiltration. Elevated cytotoxic effector molecule expression (Fas ligand, perforin, granzyme B) indicates T-cell mediated graft destruction by cytotoxic and cytolytic mechanisms within 48 to 72 h after transplantation. These results identify the CXCR3-mediated chemoattractant pathway as an immunosuppressive target in pig-to-primate islet xenotransplantation.

  • 511.
    Hårdstedt, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Center for Clinical Research Dalarna.
    Lindblom, Susanne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Hong, Jaan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Nilsson, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Ronquist, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Biochemial structure and function.
    A novel model for studies of blood-mediated long-term responses to cellular transplants2015In: Upsala Journal of Medical Sciences, ISSN 0300-9734, E-ISSN 2000-1967, Vol. 120, no 1, p. 28-39Article in journal (Refereed)
    Abstract [en]

    Aims

    Interaction between blood and bio-surfaces is important in many medical fields. With the aim of studying blood-mediated reactions to cellular transplants, we developed a whole-blood model for incubation of small volumes for up to 48 h.

    Methods

    Heparinized polyvinyl chloride tubing was cut in suitable lengths and sealed to create small bags. Multiple bags, with fresh venous blood, were incubated attached to a rotating wheel at 37°C. Physiological variables in blood were monitored: glucose, blood gases, mono- and divalent cations and chloride ions, osmolality, coagulation (platelet consumption, thrombin-antithrombin complexes (TAT)), and complement activation (C3a and SC5b-9), haemolysis, and leukocyte viability.

    Results

    Basic glucose consumption was high. Glucose depletion resulted in successive elevation of extracellular potassium, while sodium and calcium ions decreased due to inhibition of energy-requiring ion pumps. Addition of glucose improved ion balance but led to metabolic acidosis. To maintain a balanced physiological environment beyond 6 h, glucose and sodium hydrogen carbonate were added regularly based on analyses of glucose, pH, ions, and osmotic pressure. With these additives haemolysis was prevented for up to 72 h and leukocyte viability better preserved. Despite using non-heparinized blood, coagulation and complement activation were lower during long-term incubations compared with addition of thromboplastin and collagen.

    Conclusion

    A novel whole-blood model for studies of blood-mediated responses to a cellular transplant is presented allowing extended observations for up to 48 h and highlights the importance of stringent evaluations and adjustment of physiological conditions.

  • 512.
    Hårdstedt, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Center for Clinical Research Dalarna.
    Lindblom, Susanne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Karlsson-Para, Alex
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Nilsson, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Characterization of innate immunity in an extended whole blood model of human islet allotransplantation2016In: Cell Transplantation, ISSN 0963-6897, E-ISSN 1555-3892, Vol. 25, no 3, p. 503-515Article in journal (Refereed)
    Abstract [en]

    The instant blood-mediated inflammatory reaction (IBMIR) has been studied in whole blood models of human allo-islet transplantation for short periods (<6 h). Beyond this time frame the innate response to intraportally transplanted islets is less well described. A novel whole blood model was applied to study blood islet graft interactions up to 48 h. Heparinized polyvinyl chloride tubing was sealed into small bags containing venous blood together with allogeneic human islets and exocrine tissue, respectively. The bags were attached to a rotating wheel (37 degrees C). Concentrated glucose and sodium hydrogen carbonate were added every 12 h to maintain physiological limits for sustained immune cell functions. Plasma was collected at repeated time points for analyses of coagulation/complement activation and chemokine/cytokine production. Immune cell infiltration was analyzed using immunohistochemistry. Coagulation and platelet activation markers, thrombin antithrombin complex (TAT) and soluble CD40 ligand (sCD4OL) showed early high concentrations (at 6-12 h). sC5b-9 steadily increased over 48 h. At 6 h neutrophils and monocytes surrounded the clotted cellular grafts with a following massive infiltration of neutrophils. High and increasing concentrations of CXCR1/2 ligands [IL-8 and growth-regulated oncogene alpha/beta/gamma (Gro-alpha/beta/gamma)] and IL-6 were produced in response to human islets and exocrine tissue. The CCR2 ligand monocyte chemoattractant protein 1 (MCP-1) exhibited increasing concentrations in response to exocrine tissue. The CXCR3 ligand interferon-inducible T cell alpha chemoattractant (I-TAC) was produced in response to both human islets and exocrine tissue from 6 h. Monokine induced by yinterferon (Mig) and interferon gamma-induced protein 10 (IP-10) showed a later response, preferentially to exocrine tissue and with larger variations among preparations. An extended blood model of clinical islet transplantation allowed characterization of early immune activation in response to human islets and exocrine tissue. Increased production of chemokines targeting CXCR1/2, CCR2, and CXCR3 was observed, accompanied by massive intraislet neutrophil infiltration over 48 h. The model proved to be useful in exploring early blood-mediated reactions to cellular transplants and has relevance for evaluation of pharmacological interventions to prevent graft loss.

  • 513. Höglind, Ankie
    et al.
    Areström, Irene
    Ehrnfelt, Cecilia
    Masjedi, Khosro
    Zuber, Bartels
    Giavedoni, Luis
    Ahlborg, Niklas
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Mabtech, Sweden.
    Systematic evaluation of monoclonal antibodies and immunoassays for the detection of Interferon-gamma and Interleukin-2 in old and new world non-human primates2017In: JIM - Journal of Immunological Methods, ISSN 0022-1759, E-ISSN 1872-7905, Vol. 441, p. 39-48Article in journal (Refereed)
    Abstract [en]

    Non-human primates (NHP) provide important animal models for studies on immune responses to infections and vaccines. When assessing cellular immunity in NHP, cytokines are almost exclusively analyzed utilizing cross-reactive anti-human antibodies. The functionality of antibodies has to be empirically established for each assay/application as well as NHP species.

    A rational approach was employed to identify monoclonal antibodies (mAb) cross-reactive with many NHP species. Panels of new and established mAbs against human Interferon (IFN)-gamma and Interleukin (IL)-2 were assessed for reactivity with eukaryotically expressed recombinant IFN-gamma and IL-2, respectively, from Old (rhesus, cynomolgus and pigtail macaques, African green monkey, sooty mangabey and baboon) and New World NHP (Ma's night monkey, squirrel monkey and common marmoset).

    Pan-reactive mAbs, recognizing cytokines from all NHP species, were further analyzed in capture assays and flow cytometry with NHP peripheral blood mononuclear cells (PBMC). Pan-reactive mAb pairs for IFN-gamma well as IL-2 were identified and used in ELISA to measure IFN-gamma and IL-2, respectively, in Old and New World NHP PBMC supernatants. The same mAb pairs displayed high functionality in ELISpot and FluoroSpot for the measurement of antigen-specific IFN-gamma and IL-2 responses using cynomolgus PBMC Functionality of pan-reactive mAbs in flow cytometry was also verified with cynomolgus PBMC.

    The development of well-defined immunoassays functional with a panel of NHP species facilitates improved analyses of cellular immunity and enables inclusion in multiplex cytokine assays intended for a variety of NHP.

  • 514.
    Högstrand, Kari
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    Gene conversion of the mouse MHC class II1998Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The major histocompatibility complex (MHC) is the most polymorphic vertebrate gene loci known to exist. Gene products of the MHC are responsible for presentation of antigen derived peptides to T cells of the immune system, which thereby become activated and involved in the elimination of particles recognised as non-self. The mechanism of acquiring this extensive polymorphism is not known, but gene conversion has been suggested as one possible molecular genetic mechanism employed to generate new MHC alleles. A strict definition of gene conversion, based on the phenomenon in yeast, involves the transfer of a DNA fragment from a donor gene to a homologous acceptor gene without the donor gene being changed in the process. For practical reasons, the term gene conversion in higher organisms is rather used in the context of ìtemplated segmental mutationì.

    This thesis describes the first direct evidence of gene conversion events between two endogenous MHC class II genes in a mouse system using sperm as a representative for future potential individuals. A PCR assay was developed using primers for the acceptor gene as well as the donor gene to obtain a detectable product only in cells where a gene conversion event had occurred. Interchromosomal gene conversion between a MHC class II Ab and Eb gene located on separate chromosomes was found to occur at a frequency of 1/500.000 sperm, whereas no gene conversion was detected in somatic liver cells. Intrachromosomal gene conversions between Ab and Eb genes located on the same chromosome were detected at frequencies ranging from 1/35.000 sperm to 1/830.000 sperm depending on the alleles in ves gat ed. This difference in frequency was found although the investigated alleles coexisted in the same cell, which indicates that there is sequence restraints acting on the genes involved in a gene conversion event. Both inter- and intra-chromosomal gene conversions transferred DNA fragments which varied in length, but the transfer breakpoints investigated were usually in regions where the donor and acceptor genes showed a high nucleotide similarity.

    Investigation of gene conversion during male gametogenesis showed that most gene conversion events detected between MHC class II genes were completed already in the premeiotic spermatogonia cell stage, which indicates that gene conversion relies on other molecular genetic mechanisms than normal meiotic recombination. Sequence analysis of several MHC mutants proposed to have been caused by gene conversion events revealed that both donor and acceptor sequences resided in regions where normal mammalian CpG suppression was absent. CpG dinucleotides have been shown to be relatively mutation prone, which accordingly suggests that regions with a high CpG content could be targeted for mutation events. Moreover, gene conversion could be induced and increased up to 15 times of the background level in a cell line by DNA damage, which indicates that the DNA repair system is involved in the gene conversion mechanism.

    From an evolutionary point of view, gene conversion thus is a very potent mechanism for creation of new MHC alleles by shuffling of already existing and functional parts of genes into new homologous locations. This mechanism can efficiently produce new functional MHC variants, which could give the population a selective advantage in terms of the ability to cope with various pathogens as well as being a mean to avoid inbreeding that could result in a reproductive loss.

  • 515.
    Ihnatko, Robert
    et al.
    Institute of Virology, Slovak Academy of Sciences, Dúbravská cesta 9, 842 45 Bratislava, Slovakia.
    Kubes, M
    Institute of Virology, Slovak Academy of Sciences, Dúbravská cesta 9, 842 45 Bratislava, Slovakia.
    TNF signaling: early events and phosphorylation.2007In: General Physiology and Biophysics, ISSN 0231-5882, E-ISSN 1338-4325, Vol. 26, no 3, p. 159-67Article in journal (Refereed)
    Abstract [en]

    Tumor necrosis factor-alpha (TNF) is a major mediator of apoptosis as well as immunity and inflammation. Inappropriate production of TNF or sustained activation of TNF signaling has been implicated in the pathogenesis of a wide spectrum of human diseases, including cancer, osteoporosis, sepsis, diabetes, and autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, and inflammatory bowel disease. TNF binds to two specific receptors, TNF-receptor type I (TNF-R1, CD120a, p55/60) and TNF-receptor type II (TNF-R2, CD120b, p75/80). Signaling through TNF-R1 is extremely complex, leading to both cell death and survival signals. Many findings suggest an important role of phosphorylation of the TNF-R1 by number of protein kinases. Role of TNF-R2 phosphorylation on its signaling properties is understood less than TNF-R1. Other cellular substrates as TRADD adaptor protein, TRAF protein family and RIP kinases are reviewed in relation to TNF receptor-mediated apoptosis or survival pathways and regulation of their actions by phosphorylation.

  • 516.
    Ihnatko, Robert
    et al.
    Slovak Academy of Sciences, Institute of Virology, Laboratory for Diagnosis and Prevention of Rickettsial and Chlamydial Infections, Slovak Republic.
    Vadovič, Pavol
    Slovak Academy of Sciences, Institute of Virology, Laboratory for Diagnosis and Prevention of Rickettsial and Chlamydial Infections, Slovak Republic.
    Toman, Rudolf
    Slovak Academy of Sciences, Institute of Virology, Laboratory for Diagnosis and Prevention of Rickettsial and Chlamydial Infections, Slovak Republic.
    Proteins of Coxiella Burnetii and Analysis of their Function2011In: BSL3 and BSL4 Agents: Proteomics, Glycomics, and Antigenicity / [ed] Jiri Stulik, Rudolf Toman, Patrick Butaye and Robert G. Ulrich, Wiley-VCH Verlag GmbH & Co. KGaA , 2011, p. 145-151Chapter in book (Refereed)
  • 517.
    Ikebuchi, Ryoyo
    et al.
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan;Japan Soc Promot Sci, Tokyo, Japan.
    Fujimoto, Maika
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Nakanishi, Yasutaka
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Okuyama, Hiromi
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Moriya, Taiki
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Kusumoto, Yutaka
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Tomura, Michio
    Osaka Ohtani Univ, Fac Pharm, Lab Immunol, Tondabayashi, Japan.
    Functional Phenotypic Diversity of Regulatory T Cells Remaining in Inflamed Skin2019In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 10, article id 1098Article in journal (Refereed)
    Abstract [en]

    Regulatory T cells (Tregs) migrate between lymphoid and peripheral tissues for maintaining immune homeostasis. Tissue-specific function and functional heterogeneity of Tregs have been suggested, however, correlation between them and inter-tissue movement remain unknown. We used a contact hypersensitivity model of mice expressing a photoconvertible protein for tracking migratory cells. After marking cells in skin, we purified Tregs exhibiting a different migration pattern [Tregs recruiting to or remaining in the skin and emigrating from the skin to draining lymph nodes (dLNs) within half a day] and examined single-cell gene and protein expression profiles. Correlation and unsupervised clustering analyses revealed that Tregs in both skin and dLNs comprised two subpopulations, one highly expressing Nrp1 with variable CD25, Granzyme B, and/or CTLA-4 expression and another with 3 subsets strongly expressing CD25, Granzyme B, or CTLA-4 together with CD39. Characteristic subsets of Tregs remaining in the skin displayed higher CD25 and CD39 expression and lower Granzyme B and CTLA-4 expression compared with Tregs migrating to the skin. In addition, CCR5 expression in Tregs in skin was positively and negatively correlated with CD39 and Nrp-1 expression, respectively. To assess the predictive value of these data for immunotherapy, we blocked CCR5 signaling and found modest downregulation of CD39 and modest upregulation of Nrp1 expression in skin Tregs. Our data reveal a high functional diversity of Tregs in skin that is strongly related to trafficking behavior, particularly skin retention. Modulation of tissue-specific trafficking and function is a promising clinical strategy against autoimmune, infectious, and neoplastic diseases.

  • 518. Ilander, Mette
    et al.
    Olsson-Stromberg, Ulla
    Lahteenmaki, Hanna
    Kasanen, Tiina
    Koskenvesa, Perttu
    Soderlund, Stina
    Hoglund, Martin
    Markevärn, Berit
    Själander, Anders
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine, Medicine.
    Lofti, Kouros
    Malm, Claes
    Lubking, Anna
    Ekblom, Marja
    Holm, Elena
    Bjoreman, Mats
    Lehmann, Soren
    Stenke, Leif
    Ohm, Lotta
    Hjorth-Hansen, Henrik
    Saussele, Susanne
    Mahon, Francois-Xavier
    Porkka, Kimmo
    Richter, Johan
    Mustjoki, Satu
    Disease Relapse After Tyrosine Kinase Inhibitor Treatment Discontinuation in Chronic Myeloid Leukaemia is Related to Both Low Number and Impaired Function of NK Cells2014In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 79, no 6, p. 467-468Article in journal (Other academic)
  • 519. Ilander, Mette
    et al.
    Olsson-Strömberg, Ulla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Lahteenmaki, Hanna
    Kasanen, Tiina
    Koskenvesa, Perttu
    Söderlund, Stina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Höglund, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Markevarn, Berit
    Sjalander, Anders
    Lofti, Kouros
    Malm, Claes
    Lubking, Anna
    Ekblom, Marja
    Holm, Elena
    Bjoreman, Mats
    Lehmann, Soren
    Stenke, Leif
    Ohm, Lotta
    Hjorth-Hansen, Henrik
    Saussele, Susanne
    Mahon, Francois-Xavier
    Porkka, Kimmo
    Richter, Johan
    Mustjoki, Satu
    Disease Relapse After Tyrosine Kinase Inhibitor Treatment Discontinuation in Chronic Myeloid Leukaemia is Related to Both Low Number and Impaired Function of NK Cells2014In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 79, no 6, p. 467-468Article in journal (Other academic)
  • 520.
    Imgenberg-Kreuz, Juliana
    et al.
    Uppsala Univ, Sweden.
    Carlsson Almlöf, Jonas Carlsson
    Uppsala Univ, Sweden.
    Leonard, Dag
    Uppsala Univ, Sweden.
    Sjöwall, Christopher
    Linköping University, Department of Clinical and Experimental Medicine, Division of Neuro and Inflammation Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Rheumatology.
    Syvanen, Ann-Christine
    Uppsala Univ, Sweden.
    Ronnblom, Lars
    Uppsala Univ, Sweden.
    Sandling, Johanna K.
    Uppsala Univ, Sweden.
    Nordmark, Gunnel
    Uppsala Univ, Sweden.
    Shared and Unique Patterns of DNA Methylation in Systemic Lupus Erythematosus and Primary Sjogrens Syndrome2019In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 10, article id 1686Article in journal (Refereed)
    Abstract [en]

    Objectives: To performa cross-comparative analysis of DNA methylation in patients with systemic lupus erythematosus (SLE), patients with primary Sjogrens syndrome (pSS), and healthy controls addressing the question of epigenetic sharing and aiming to detect disease-specific alterations. Methods: DNA extracted from peripheral blood from 347 cases with SLE, 100 cases with pSS, and 400 healthy controls were analyzed on the Human Methylation 450k array, targeting 485,000 CpG sites across the genome. A linear regression model including age, sex, and blood cell type distribution as covariates was fitted, and association p-values were Bonferroni corrected. A random forest machine learning classifier was designed for prediction of disease status based on DNA methylation data. Results: We established a combined set of 4,945 shared differentially methylated CpG sites (DMCs) in SLE and pSS compared to controls. In pSS, hypomethylation at type I interferon induced genes was mainly driven by patients who were positive for Ro/SSA and/or La/SSB autoantibodies. Analysis of differential methylation between SLE and pSS identified 2,244 DMCs with a majority of sites showing decreased methylation in SLE compared to pSS. The random forest classifier demonstrated good performance in discerning between disease status with an area under the curve (AUC) between 0.83 and 0.96. Conclusions: The majority of differential DNA methylation is shared between SLE and pSS, however, important quantitative differences exist. Our data highlight neutrophil dysregulation as a shared mechanism, emphasizing the role of neutrophils in the pathogenesis of systemic autoimmune diseases. The current study provides evidence for genes and molecular pathways driving common and disease-specific pathogenic mechanisms.

  • 521.
    Imgenberg-Kreuz, Juliana
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Sandling, Johanna K.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Bjork, A.
    Karolinska Inst, Dept Med, Karolinska Univ Hosp, Stockholm, Sweden.
    Nordlund, J.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Kvarnstrom, M.
    Karolinska Inst, Dept Med, Karolinska Univ Hosp, Stockholm, Sweden.
    Eloranta, Maija-Leena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Rönnblom, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Wahren-Herlenius, M.
    Karolinska Inst, Dept Med, Karolinska Univ Hosp, Stockholm, Sweden.
    Syvänen, Ann-Christine
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Nordmark, Gunnel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Transcription profiling of peripheral B cells in antibody-positive primary Sjogren's syndrome reveals upregulated expression of CX3CR1 and a type I and type II interferon signature2018In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 87, no 5, article id UNSP e12662Article in journal (Refereed)
    Abstract [en]

    B cells play a key role in the pathogenesis of primary Sjogren's syndrome (pSS). The aim of this study was to analyse the transcriptome of CD19+ B cells from patients with pSS and healthy controls to decipher the B cell-specific contribution to pSS. RNA from purified CD19+ B cells from 12 anti-SSA antibody-positive untreated female patients with pSS and 20 healthy blood donors was subjected to whole transcriptome sequencing. A false discovery rate corrected significance threshold of <0.05 was applied to define differential gene expression. As validation, gene expression in B cells from 17 patients with pSS and 16 healthy controls was analysed using a targeted gene panel. RNA-sequencing identified 4047 differentially expressed autosomal genes in pSS B cells. Upregulated expression of type I and type II interferon (IFN)-induced genes was observed, establishing an IFN signature in pSS B cells. Among the top upregulated and validated genes were CX3CR1, encoding the fractalkine receptor involved in regulation of B-cell malignancies, CCL5/RANTES and CCR1. Increased expression of several members of the TNF superfamily was also identified; TNFSF4/Ox40L, TNFSF10/TRAIL, TNFSF13B/BAFF, TNFRSF17/BCMA as well as S100A8 and -A9/calprotectin, TLR7, STAT1 and STAT2. Among genes with downregulated expression in pSS B cells were SOCS1 and SOCS3, CD70 and TNFAIP3/A20. We conclude that B cells from patients with anti-SSA antibody-positive pSS display immune activation with upregulated expression of chemokines, chemokine receptors and a prominent type I and type II IFN signature, while suppressors of cytokine signalling are downregulated. This adds insight into the autoimmune process and suggests potential targets for future functional studies.

  • 522.
    Imgenberg-Kreuz, Juliana
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Sandling, Johanna K.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Nordmark, Gunnel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Epigenetic alterations in primary Sjogren's syndrome: an overview2018In: Clinical Immunology, ISSN 1521-6616, E-ISSN 1521-7035, Vol. 196, p. 12-20Article in journal (Refereed)
    Abstract [en]

    Primary Sjogren's syndrome (pSS) is a chronic autoimmune rheumatic disease characterized by inflammation of exocrine glands, mainly salivary and lacrimal glands. In addition, pSS may affect multiple other organs resulting in systemic manifestations. Although the precise etiology of pSS remains elusive, pSS is considered to be a multi factorial disease, where underlying genetic predisposition, environmental factors and epigenetic mechanisms contribute to disease development. Epigenetic mechanisms, such as DNA methylation, histone modifications and non-coding RNAs, may constitute a dynamic link between genome, environment and phenotypic manifestation by their modulating effects on gene expression. A growing body of studies reporting altered epigenetic landscapes in pSS suggests that epigenetic mechanisms play a role in the pathogenesis of pSS, and the reversible nature of epigenetic modifications suggests therapeutic strategies targeting epigenetic dysregulation in pSS. This article reviews our current understanding of epigenetic mechanisms in pSS and discusses implications for novel diagnostic and therapeutic approaches.

  • 523.
    Ingelsson, Björn
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Söderberg, Daniel
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences.
    Strid, Tobias
    Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Söderberg, Anita
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Bergh, Ann-Charlotte
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Loitto, Vesa-Matti
    Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Lotfi, Kourosh
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Surgery, Orthopaedics and Cancer Treatment, Department of Haematology.
    Segelmark, Mårten
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Heart and Medicine Center, Department of Nephrology.
    Spyrou, Giannis
    Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Rosén, Anders
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences.
    Lymphocytes eject interferogenic mitochondrial DNA webs in response to CpG and non-CpG oligodeoxynucleotides of class C2018In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 115, no 3, p. E478-E487Article in journal (Refereed)
    Abstract [en]

    Circulating mitochondrial DNA (mtDNA) is receiving increasing attention as a danger-associated molecular pattern in conditions such as autoimmunity, cancer, and trauma. We report here that human lymphocytes [B cells, T cells, natural killer (NK) cells], monocytes, and neutrophils derived from healthy blood donors, as well as B cells from chronic lymphocytic leukemia patients, rapidly eject mtDNA as web filament structures upon recognition of CpG and non-CpG oligodeoxynucleotides of class C. The release was quenched by ZnCl2, independent of cell death (apoptosis, necrosis, necroptosis, autophagy), and continued in the presence of TLR9 signaling inhibitors. B-cell mtDNA webs were distinct from neutrophil extracellular traps concerning structure, reactive oxygen species (ROS) dependence, and were devoid of antibacterial proteins. mtDNA webs acted as rapid (within minutes) messengers, priming antiviral type I IFN production. In summary, our findings point at a previously unrecognized role for lymphocytes in antimicrobial defense, utilizing mtDNA webs as signals in synergy with cytokines and natural antibodies, and cast light on the interplay between mitochondria and the immune system.

  • 524. Ingelsten, Madeleine
    et al.
    Gustafsson, Karin
    Olausson, Michael
    Haraldsson, Borje
    Karlsson-Parra, Alex
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Nyström, Jenny
    Rapid Increase of Interleukin-10 Plasma Levels After Combined Auxiliary Liver-Kidney Transplantation in Presensitized Patients2014In: Transplantation, ISSN 0041-1337, E-ISSN 1534-6080, Vol. 98, no 2, p. 208-215Article in journal (Refereed)
    Abstract [en]

    Background. After transplantation, donor dendritic cells (DCs) in the grafted organ are activated by an ischemia/reperfusion-induced inflammatory process that induces their migration to the recipient's secondary lymphoid tissues. The subsequent interaction between migrated and mature donor DCs, recipient T cells, and natural killer (NK) cells is proposed to be crucial in directing host immune reactions toward allograft rejection. A liver transplant is less prone to induce rejection compared with most other solid organ transplants, and simultaneous transplantation of liver and kidney is known to improve the clinical outcome of kidney transplantation. Methods and Results. Here we show that liver as well as combined auxiliary liver-kidney transplantation in patients induces a rapid increase in plasma interleukin-10 (IL-10) to levels that are significantly higher than those seen after standard kidney transplantation. Addition of IL-10 during in vitro maturation of human monocyte-derived DCs with ischemia/reperfusion-associated factors was found to affect phenotypic DC maturation significantly. Addition of IL-10 inhibited DC production of the NK cell- and T cell-recruiting chemokines CXCL9, CXCL10 and CXCL11. Conclusion. Our findings indicate that liver transplantation induces a substantial systemic release of IL-10, which may inhibit T cell- and NK cell- mediated rejection processes toward the transplanted liver and concurrently transplanted kidney.

  • 525.
    Ingemansson, Sofie
    et al.
    Lund University.
    Vaziri-Sani, Fariba
    Lund University.
    Lindblad, Ulf
    University of Gothenburg.
    Gudbjornsdottir, Soffia
    The Nordic Research Academy for Global Health.
    Törn, Carina
    Lund University.
    Long-term sustained autoimmune response to beta cell specific zinc transporter (ZnT8, W, R, Q) in young adult patients with preserved beta cell function at diagnosis of diabetes2013In: Autoimmunity, ISSN 0891-6934, E-ISSN 1607-842X, Vol. 46, no 1, p. 50-61Article in journal (Refereed)
    Abstract [en]

    The aim of this study was to examine whether autoantibodies to: ZnT8-Tryptophan (ZnT8WA), ZnT8-Arginine (ZnT8RA) or ZnT8-Glutamine (ZnT8QA) correlated with C-peptide or other autoantibodies and to assess diagnostic sensitivity of ZnT8WRQA. Specimens from 270 newly diagnosed diabetic subjects (age 15-34 years) and after 5 years duration of disease were examined. Four linear regression models were used to dissect the importance of different factors from diagnosis for the respective difference of (logZnT8WA), (logZnT8RA) and (logZnT8QA); A) unadjusted model for: initial C-peptide, age, BMI, gender, clinical classification, ICA, GADA, IA-2A, (ZnT8WA/ZnT8RA/ZnT8QA); B) C-peptide corrected for clinical factors; C) C-peptide corrected for autoantibodies; D) C-peptide corrected for all factors. The least decrease of ZnT8WA was observed in patients with high initial C-peptide in all models A) p = 0.054; B) p = 0.021; C) p = 0.047 and D) p = 0.017. A less statistically significant decrease of ZnT8RA was observed in patients with high initial C-peptide in A) p = 0.038 and C) p = 0.047, but this finding was not confirmed in B or D. The decrease of ZnT8QA levels was not related to C-peptide in any model but correlated to age D) p = 0.049. Furthermore, patients with unclassifiable diabetes showed the least decrease in D) p = 0.035. ZnT8WA, ZnT8RA or ZnT8QA were identified as a single autoantibody in 3.8% (10/266) of patients, thereby increasing diagnostic sensitivity from 79.3% (211/266) to 83.1% (221/266). In conclusion, high initial C-peptide was the most important factor even after adjusting for other factors in patients positive for ZnT8WA or ZnT8RA to remain autoantibody positive 5 years after diagnosis.

  • 526.
    Iriemenam, Nnaemeka C.
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    Antibody responses and Fc gamma receptor IIa polymorphism in relation to Plasmodium falciparum malaria2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Immunity to asexual blood-stage of Plasmodium falciparum malaria is believed to be associated with protective antibodies of certain immunoglobulin classes and subclasses. This thesis addressed the importance of antibodies in relation to malaria infection and their effective interactions with Fc gamma receptor IIa (FcyRIIa) polymorphisms. Our data indicate that the frequency of FcyRIIa-R/R131 genotype was statistically significantly higher in Sudanese patients with severe malaria, while the FcyRIIa-H/H131 genotype showed a significant association with mild malaria. The levels of IgG1 and IgG3 subclass antibodies were statistically higher in the mild malaria patients.

    The Fulani ethnic group in West Africa has been shown to be relatively resistant to malaria. We investigated the possible impact of FcyRIIa polymorphisms in the Fulani and non-Fulani in Mali and Sudan, and analysed their malaria-reactive IgG subclass profiles. The FcyRIIa-H/H131 genotype and H131-allele were found to be prevalent in the Fulani while R131-allele was prevalent in non-Fulani. The Fulani had higher serum levels of IgG1-3, with higher proportion of IgG2 than the non-Fulani.

    Most clinico-epidemiology studies have been in areas with holo- and hyper-malaria endemicity. We have analysed antibody responses to a panel of six blood-stage antigens in relation to clinical malaria outcome in mesoendemic Sudan. Our results revealed a linear association with anti-AMA-1 IgG1 antibodies and reduced risk of severe malaria while a non-linear relationship with IgG3 antibodies was observed for MSP-2, MSP-3 and GLURP. In the combined final model, the highest levels of IgG1 subclass antibodies to AMA-1, GLURP-R0, and the highest levels of IgG3 subclass antibodies reactive to 3D7 MSP-2 were independently associated with a reduced risk of clinical malaria.

    Taken together, these data suggest a possible association between FcyRIIa-R/H131 and anti-malarial antibody responses in the clinical outcome of malaria.

     

  • 527.
    Iriemenam, Nnaemeka C.
    et al.
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    Khirelsied, Atif H.
    Nasr, Amre
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    ElGhazali, Gehad
    Giha, Haider A.
    Elhassan A-Elgadir, Thoraya
    Agab-Aldour, Ahmed A.
    Montgomery, Scott M.
    Anders, Robin F.
    Theisen, Michael
    Troye-Blomberg, Marita
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    Elbashir, Mustafa I.
    Berzins, Klavs
    Stockholm University, Faculty of Science, The Wenner-Gren Institute .
    Antibody responses to a panel of Plasmodium falciparum malaria blood-stage antigens in relation to clinical disease outcome in Sudan2009In: Vaccine, ISSN 0264-410X, E-ISSN 1873-2518, Vol. 27, no 1, p. 62-71Article in journal (Refereed)
    Abstract [en]

    Despite many intervention programmes aimed at curtailing the scourge, malaria remains a formidable problem of human health. Immunity to asexual blood-stage of Plasmodium falciparum malaria is thought to be associated with protective antibodies of certain immunoglobulin classes and subclasses. We have analysed immunoglobulin G profiles to six leading blood-stage antigens in relation to clinical malaria outcome in a hospital-based study in Sudan. Our results revealed a linear association with anti-AMA-1-IgG1 antibodies in children <5 years and reduced risk of severe malaria, while the responses of the IgG3 antibodies against MSP-2, MSP-3, GLURP in individuals above 5 years were bi-modal. A dominance of IgG3 antibodies in >5 years was also observed. In the final combined model, the highest levels of IgG1 antibodies to AMA-1, GLURP-R0, and the highest levels of IgG3 antibodies to 3D7 MSP-2 were independently associated with protection from clinical malaria. The study provides further support for the potential importance of the studied merozoite vaccine candidate antigens as targets for parasite neutralizing antibody responses of the IgG1 and IgG3 subclasses.

  • 528. Isaksson, H. S.
    et al.
    Farkas, S. A.
    Mueller, P.
    Gustafsson, D.
    Nilsson, Torbjörn K.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Clinical chemistry.
    Whole genome microarray expression analysis in blood identifies pathways linked to signs and symptoms of a patient with hypercalprotectinaemia and hyperzincaemia2018In: Clinical and Experimental Immunology, ISSN 0009-9104, E-ISSN 1365-2249, Vol. 191, no 2, p. 240-251Article in journal (Refereed)
    Abstract [en]

    A child, 2 years with the hypercalprotectinaemia with hyperzincaemia' clinical syndrome, presented with atypical symptoms and signs, notably persistent fever of approximately 38 degrees C, thrombocythaemia of >700x10(9)/l and a predominance of persistent intestinal symptoms. In an effort to find a cure by identifying the dysregulated pathways we analysed whole-genome mRNA expression by the Affymetrix HG U133 Plus 20 array in blood on three occasions 3-5 months apart. Major up-regulation was demonstrated for the Janus kinase/signal transducer and activators of transcription (JAK/STAT) pathway including, in particular, CD177, S100A8, S100A9 and S100A12, accounting for the thrombocytosis; a large number of interleukins, their receptors and activators, accounting for the febrile apathic state; and the high mobility group box 1 (HMBG1) gene, possibly accounting for part of the intestinal symptoms. These results show that gene expression array technology may assist the clinician in the diagnostic work-up of individual patients with suspected syndromal states of unknown origin, and the expression data can guide the selection of optimal treatment directed at the identified target pathways.

  • 529.
    Isaksson, Helena S.
    et al.
    Örebro University, School of Medical Sciences. Department of Laboratory Medicine, Faculty of Health and Medical Sciences, Örebro University, Örebro, Sweden.
    Farkas, Sanja A.
    Örebro University, School of Health Sciences. Örebro University Hospital. Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Müller, Patrick
    Affymetrix core facility at Novum, BEA, Karolinska Institute, Huddinge, Sweden.
    Gustafsson, Dan
    Department of Pediatrics, Örebro University Hospital, Örebro, Sweden; Faculty of Health and Medical Sciences, Örebro University, Örebro, Sweden.
    Nilsson, Torbjörn, K.
    Department of Medical Biosciences, Clinical Chemistry, Umeå University, Umeå, Sweden.
    Whole genome microarray expression analysis in blood leucocytes identifies pathways linked to signs and symptoms of a patient with hypercalprotectinaemia and hyperzincaemia2018In: Clinical and Experimental Immunology, ISSN 0009-9104, E-ISSN 1365-2249, Vol. 191, no 2, p. 240-251Article in journal (Refereed)
    Abstract [en]

    A child, 2 years with the "hypercalprotectinemia with hyperzincemia" clinical syndrome presented with atypical symptoms and signs, notably persistent fever of around 38°C, thrombocythaemia of >700 x 10(9) /L, and a predominance of persistent intestinal symptoms. In an effort to find a cure by identifying the dysregulated pathways we analyzed whole-genome mRNA expression by the Affymetrix HG U133 PLUS 2.0 array on three occasions 3 to 5 months apart. Major upregulation was demonstrated for the JAK/STAT pathway including in particular CD177, S100A8, S100A9, and S100A12, accounting for the thrombocytosis; a large number of interleukins, their receptors, and activators, accounting for the febrile apathic state; and the HMBG1 gene, possibly accounting for part of the intestinal symptoms. These results show that gene expression array technology may assist the clinician in the diagnostic workup of individual patients with suspected syndromal states of unknown origin, and the expression data can guide the selection of optimal treatment directed at the identified target pathways.

  • 530.
    Isaksson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine.
    Initiation of Autoimmunity in Experimental Autoimmune Encephalomyelitis2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The events that trigger an autoimmune disease remain largely unknown. To study these events animal models are necessary because symptoms of autoimmune diseases are preceded by a long asymptomatic period in humans.

    Experimental autoimmune encephalomyelitis (EAE) is the best characterized model for cell mediated autoimmunity and an animal model for the human disease multiple sclerosis. EAE is induced in rodents by immunization with myelin antigens (Ags) together with adjuvants. After immunization, T cells are primed in the periphery by Ag presenting cells and subsequently invade the central nervous system where they mediate parenchymal inflammation, resulting in demyelination and clinical symptoms of an ascending paralysis. It is now generally recognised that the main cell type mediating EAE is the T helper type 17 (Th17) cell.

    Tolerance to EAE can be attained by DNA vaccination, but how the immune response against the myelin Ags is abrogated after DNA vaccination is not known. By employing short interfering RNA technology, induction of the innate immune signalling molecule interferon (IFN) -β was found to be necessary for the protective effect of DNA vaccination in EAE. In addition, DNA vaccination inhibited subsequent autoimmune Th17 cell responses.

    The Toll-like receptors (TLRs) of the innate immune system have evolved to recognise conserved molecular structures on microbes and signalling through them almost exclusively converge on the molecule MyD88. Signalling via MyD88 was found to be required for induction of EAE since mice deficient in this molecule did not develop disease. Upstream signalling via TLR4 and TLR9 had tolerogenic properties.

    In studies of Ag presentation in EAE, two major subtypes of dendritic cells (DCs) were examined. Plasmacytoid DCs were found to have a promoting role in the induction of EAE, partly via type 1 IFNs. Myeloid DCs had a redundant role in the induction phase of EAE, neither disease severity nor encephalitogenic Th17 responses were affected by their absence during priming.

    These studies further demonstrate that the cells and molecules of the innate immune system exhibit a crucial role in controlling the adaptive immune system which mediates tissue damage in autoimmune diseases.

  • 531.
    Isaksson, Magnus
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Autoimmunity.
    Ardesjö Lundgren, Brita
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Autoimmunity.
    Ahlgren, Kerstin M
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Autoimmunity.
    Kämpe, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Autoimmunity.
    Lobell, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Autoimmunity.
    Conditional DC depletion does not affect priming of encephalitogenic Th cells in EAE2012In: European Journal of Immunology, ISSN 0014-2980, E-ISSN 1521-4141, Vol. 42, no 10, p. 2555-2563Article in journal (Refereed)
    Abstract [en]

    EAE, an animal model for multiple sclerosis, is a Th17- and Th1-cell-mediated auto-immune disease, but the mechanisms leading to priming of encephalitogenicTcells in autoimmune neuroinflammation are poorly understood. To investigate the role of dendritic cells (DCs) in the initiation of autoimmuneTh17- andTh1-cell responses andEAE, we used mice transgenic for a simian diphtheria toxin receptor (DTR) expressed under the control of the murineCD11c promoter (CD11c-DTRmice onC57BL/6 background).EAEwas induced by immunization with myelin oligodendrocyte glycoprotein (MOG) protein in CFA. DCs were depleted on the day before and 8 days afterMOG immunization. The mean clinicalEAEscore was only mildly reduced inDC-depleted mice when DCs were ablated beforeEAEinduction. The frequency of activatedTh cells was not altered, andMOG-inducedTh17 orTh1-cell responses were not altered, in the spleens ofDC-depleted mice. Similar results were obtained ifDCswere ablated the first 10 days afterMOGimmunization with repeatedDCdepletions. Unexpectedly, transient depletion of DCs did not affect priming or differentiation of MOG-inducedTh17 andTh1-cell responses or the incidence ofEAE. Thus, the mechansim of priming ofTh cells inEAEremains to be elucidated.

  • 532.
    Israelsson, Elisabeth
    Stockholm University, Faculty of Science, Wenner-Gren Institute for Experimental Biology.
    Host genetic factors and antibody responses with potential involvement in the susceptibility to malaria2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The relatively lower susceptibility to malaria seen in the Fulani ethnic group in Africa, as compared to other sympatric ethnic groups, has been related to genetic regulation of the immune responses. This thesis aimed to describe important pathways related to the regulation of antibodies in the immune responses during a malaria infection. Our results suggest that the higher anti-malarial immune responses seen in the Fulani are not a general hyper-responsiveness in this group, but neither a malaria specific response. Fcγ receptors are important structures in the immune responses, and polymorphisms in these genes were associated with IgG subclass levels, P. falciparum parasitemia and haemoglobin levels, suggesting that these polymorphisms may be a contributing factor to the differential susceptibility to malaria. C-reactive protein levels rise immediately in response to inflammatory stimuli, and the -286 CRP polymorphism was indicated to influence parasite levels, suggesting a possible involvement in the lower susceptibility to malaria seen in the Fulani ethnic group. Several cytokines are important in maintaining the optimal parasite-neutralizing milieu in the host, and we investigated polymorphisms in some of these cytokine genes, in order to establish a possible influence of these on malaria susceptibility. Several of these polymorphisms showed associations with haemoglobin levels, IgG subclass antibody levels and parasitemia, suggesting that IL-1β, IL-6, IL-10 and TNF could affect the susceptibility to malaria and the severity of the malaria infection.

    Taken together, these data suggest that genetic factors have the ability to affect the antibody responses, and that several pathways can be affected. Moreover, the Fulani have a genetic predisposition for a higher inflammatory response during a malaria infection, which could lower their susceptibility to the disease. However, the control measures for this inflammation still have to be established and evaluated.

  • 533.
    Israelsson, Elisabeth
    et al.
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Ekström, Mattias
    Nasr, Amre
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Dolo, Amagana
    Kearsley, Susannah
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Arambepola, Gishanthi
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Homann, Manijeh V.
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Maiga, Bakary
    Doumbo, Ogobara K.
    ElGhazali, Gehad
    Giha, Hayder A.
    Troye-Blomberg, Marita
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Berzins, Klavs
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Tornvall, Per
    Marked differences in CRP genotype frequencies between the Fulani and sympatric ethnic groups in Africa2009In: Malaria Journal, ISSN 1475-2875, E-ISSN 1475-2875, Vol. 8, no 136Article in journal (Refereed)
    Abstract [en]

    Background

    C-reactive protein (CRP) is an acute phase protein that can activate various immune cells and bind to certain Fcγ receptors. The latter may compete with the binding of IgG antibodies to these receptors and could thereby interfere with the antigen-specific immune response. Polymorphisms in the promoter region of the CRP gene have been strongly associated with the plasma concentration of CRP. The known lower susceptibility to malaria in the Fulani ethnic group, as compared to their sympatric neighbours in Africa, has been linked to different genetic backgrounds. The present study was performed to investigate if polymorphisms in the CRP gene could contribute to the lower susceptibility to malaria seen in the Fulani ethnic group.

    Methods

    The CRP -717 T>C, -286 C>T>A, and +1444 C>T polymorphisms were analysed in asymptomatic Fulani and non-Fulani individuals from Mali and Sudan using Pyrosequencing T and TaqMan r MGB probes.

    Results

    The rare -286 A allele, previously shown to be associated with increased CRP expression and plasma levels, was shown to be more frequent in the non-Fulani ethnic groups as compared to the sympatric Fulani ethnic group both in Mali and Sudan. The common -717 T allele was more prevalent in the non-Fulani ethnic group compared to the sympatric Fulani ethnic group, but only in Mali. The parasite prevalence was increased for the -286 A allele, but not for the -717 T allele. No differences regarding genotype frequency or parasite prevalence were seen for +1444 C>T.

    Conclusion

    This study indicate that CRP may play an important role in the immune responses to malaria, and that the -286 C/T/A CRP polymorphism may be a contributing factor to the lower susceptibility to malaria seen in the Fulani.

  • 534.
    Israelsson, Pernilla
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology. Umeå University, Faculty of Medicine, Department of Clinical Sciences, Obstetrics and Gynecology.
    Labani-Motlagh, Alireza
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Nagaev, Ivan
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Dehlin, Eva
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Nagaeva, Olga
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Lundin, Eva
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Ottander, Ulrika
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Obstetrics and Gynecology.
    Mincheva-Nilsson, Lucia
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Assessment of cytokine mRNA expression profiles in tumor microenvironment and peripheral blood mononuclear cells of patients with high-grade serous carcinoma of the ovary2017In: Journal of Cancer Science & Therapy, ISSN 1948-5956, Vol. 9, no 5, p. 422-429Article in journal (Refereed)
    Abstract [en]

    Objective: Tumor establishment, metastatic spreading and poor survival in ovarian cancer is strongly associated with progressive derangement of the patient’s immune system. Accumulating evidence suggests that immune impairment is influenced by the production and presence of cytokines in the tumor microenvironment. Methods: Cytokine mRNA profiles in tumor tissue and peripheral blood mononuclear cells (PBMC) were analyzed in patients with high grade serous carcinoma (HGSC) of the ovary and compared it to patients with benign ovarian conditions and controls with normal ovaries. Cytokine assessment was done by real-time quantitative RT-PCR and specific primers and probes for 12 cytokines-IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-15, TNF-α, TNF-β/LTA, TGF-β1, and GM-CSF chosen to distinguish between cytotoxic Th1, humoral Th2, regulatory Th3/Tr1 and inflammatory responses. Results: The cytokine mRNA response in the HGSC patients was significantly up regulated compared to patients with benign ovarian conditions and normal ovary controls confirming the immunogenicity of HGSC and implying immune recognition and reaction locally in the tumor microenvironment and systemically in the peripheral blood.There was an up-regulation of inflammatory and inhibitory cytokine mRNA promoting tumor progression, T-regulatory cell priming and T-regulatory cell-mediated immune suppression. In contrast, there was an inability to mount the crucially important IFN gamma response needed for upregulation of the cytotoxic anti-tumor response in the local microenvironment. In addition, systemic IL-4- mediated Th2 response prevailed in the peripheral blood deviating the systemic defense towards humoral immunity. Conclusions: Taken together, these results suggest local and systemic cytokine cooperation promoting tumor survival, progression and immune escape. Our study confirms and extends previous investigations and contributes to the evaluation of potential cytokine candidates for diagnostic cytokine mRNA profiles and for future therapeutic interventions based on cytokine inhibition.

  • 535. Ivanchenko, Margarita
    et al.
    Brauner, Susanna
    Enblad, Gunilla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Oncology.
    Sundström, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Backlin, Carin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Baecklund, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Wahren-Herlenius, Marie
    Ro52/TRIM21 Expression is Decreased in Malignant B Cells2014In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 79, no 6, p. 453-454Article in journal (Other academic)
  • 536.
    Ivanoff, Jyrki
    Umeå University, Faculty of Medicine, Clinical Microbiology.
    Migration on extracellular matrix surface and infiltration into matrix - two distinguishable activities of human T cells2003Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Migration of T-lymphocytes on a surface coated with extracellular matrix (ECM) components (two-dimensional (2-D) migration) and migration (infiltration) into a matrix (Three-dimesional (3-D) migration) are complex events and the underlying mechanisms are not yet fully understood. Here 2-D and 3-D migration were studied by use of seven leukemic T-cell lines representing discrete differentiation stages, a non-leukemic T-cell clone, and normal peripheral blood T cells. peripheral blood lymphocytes and the T-cell clone produced nanogram quantities of various chemokines, as compared to a production of ≤ 0.05 ng/ml by the T leukemia cell lines. In a Boyden chamber system, the leukemic T-cell lines showed haptotactic migration on fibronectin. The migration was augmented bu exposure to chemokines, including RANTES, MIP-1α, MIP-1β, and IL-8. The T-cell lines showed a peak response at a chemokine concentration of 10-50 ng/ml, whereas the T-cell clone responded optimally at 100 ng/ml. In contrast to a general capability of T-cells to migrate on 2-D ECM, only some of the T-cell lines were capable of 3-D migration into Matrigel or a collagen matrix. The infiltrative capacity was unrelated to the capacity to migrate on or adhere to the substrata. T-cell lines with a capacity to infiltrate produced matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), whereas non-infiltrating cell lines did not produce MMP-9. T-cell lines capable of infiltrating Matrigel or collagen responded to chemokines exposure with increased infiltration, but the chemokines did not render non-infiltrative cell lines infiltrative. Stimulation of infiltration of T-cell lines into collagen by the chemokine SDF-1α was inhibited by somatostatin, a neuropeptide with immunosuppressive properties. In conclusion, the ability to migrate on 2-D substrata and to infiltrate into 3.D substrata was found to be distinguishable properties of T cells. failure of some T-cell lines to infiltrate correlated with the lack of expression of MMP-9. Chemokines stimulated infiltration of infiltrative T-cell lines into collagen and Matrigel but did not render non-infiltrative T-cell lines infiltrative. Finally, a possible physiological mechanism for modulation of the chemokine-stimulated 3-D migration was demonstrated.

  • 537.
    Jacobsen, Freja A.
    et al.
    Dept. of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark & Novo Nordisk A/S, Gentofte, Denmark.
    Hulst, Camilla
    Dept. of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark & Novo Nordisk A/S, Gentofte, Denmark.
    Bäckström, Thomas
    Novo Nordisk A/S, Måløv, Denmark & BTB Pharma, Malmö, Sweden.
    Koleske, Anthony J.
    Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, USA.
    Andersson, Åsa
    Dept. of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark.
    Arg Deficiency Does not Influence the Course of Myelin Oligodendrocyte Glycoprotein (MOG35-55)-induced Experimental Autoimmune Encephalomyelitis2016In: Journal of Clinical & Cellular Immunology, ISSN 2155-9899, E-ISSN 2155-9899, Vol. 7, no 3, article id 1000420Article in journal (Refereed)
    Abstract [en]

    Background: Inhibition of Abl kinases has an ameliorating effect on the rodent model for multiple sclerosis, experimental autoimmune encephalomyelitis, and arrests lymphocyte activation. The family of Abl kinases consists of the Abl1/Abl and Abl2/Arg tyrosine kinases. While the Abl kinase has been extensively studied in immune activation, roles for Arg are incompletely characterized. To investigate the role for Arg in experimental autoimmune encephalomyelitis, we studied disease development in Arg-/- mice.

    Methods: Arg-/- and Arg+/+ mice were generated from breeding of Arg+/- mice on the C57BL/6 background. Mice were immunized with the myelin oligodendrocyte glycoprotein (MOG)35-55 peptide and disease development recorded. Lymphocyte phenotypes of wild type Arg+/+ and Arg-/- mice were studied by in vitro stimulation assays and flow cytometry.

    Results: The breeding of Arg+/+ and Arg-/- mice showed skewing in the frequency of born Arg-/- mice. Loss of Arg function did not affect development of experimental autoimmune encephalomyelitis, but reduced the number of splenic B-cells in Arg-/- mice following immunization with MOG peptide.

    Conclusions: Development of MOG-induced experimental autoimmune encephalomyelitis is not dependent on Arg, but Arg plays a role for the number of B cells in immunized mice. This might suggest a novel role for the Arg kinase in B-cell trafficking or regulation. Furthermore, the results suggest that Arg is important for normal embryonic development. © 2016 Jacobsen FA, et al.

  • 538.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Boiko, Iryna
    Clinical Laboratory Department, Ternopil Regional Clinical Dermatovenerologic Dispensary, Ternopil, Ukraine.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Blondeel, Karel
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Kiarie, James
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Toskin, Igor
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Peeling, Rosanna W.
    London School of Hygiene and Tropical Medicine (LSHTM), London, UK.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    WHO laboratory validation of Xpert((R)) CT/NG and Xpert((R)) TV on the GeneXpert system verifies high performances2018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 12, p. 907-912Article in journal (Refereed)
    Abstract [en]

    Effective tests for diagnosis of sexually transmitted infections (STIs), used point of care to inform treatment and management decisions, are urgently needed. We evaluated the analytical sensitivity and specificity of the Xpert((R)) CT/NG and Xpert((R)) TV tests, examining 339 samples spiked with phenotypically and/or genetically diverse strains of Neisseria gonorrhoeae, Chlamydia trachomatis, and Trichomonas vaginalis, and other related species that may cross-react. The APTIMA Combo 2 test and APTIMA TV test were used as reference tests. The analytical sensitivity for all three agents in the Xpert((R)) CT/NG and Xpert((R)) TV tests was <= 10(2) genome equivalents/reaction. The analytical specificity of both tests was high. False-positive results were identified in the Xpert((R)) TV test when challenging with high concentrations of Trichomonas tenax, Trichomonas gallinae, Trichomonas stableri, and Trichomonas aotus. However, the clinical relevance of these cross-reactions can likely be neglected, because these species have not been identified in urogenital samples from humans. In conclusion, the analytical sensitivity and specificity of the user-friendly Xpert((R)) CT/NG and Xpert((R)) TV tests on the GeneXpert system were high. The results support the use of specimens from also extra-genital sites, for example, pharynx and rectum. However, appropriate clinical validations are additionally required.

  • 539.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Neisseria meningitidis, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Stenmark, Bianca
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Neisseria meningitidis, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Hedberg, Sara Thulin
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Mölling, Paula
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Fredlund, Hans
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Neisseria meningitidis carriage in Swedish teenagers associated with the serogroup W outbreak at the World Scout Jamboree, Japan 20152018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 4, p. 337-341Article in journal (Refereed)
    Abstract [en]

    The aims of the study were to estimate the carrier state of Neisseria meningitidis in Swedish teenagers and its association with an outbreak at the World Scout Jamboree in 2015 as well as to compare sensitivity of throat versus nasopharyngeal swab for optimal detection of carriage. In total, 1 705 samples (cultures n = 32, throat swabs n = 715, nasopharyngeal swabs n = 958) from 1 020 Jamboree participants were collected and sent to the National Reference Laboratory for Neisseria meningitidis for culture and molecular analysis. The overall positivity for N. meningitidis was 8% (83/1 020), whereas 2% (n = 22) belonged to a known sero/genogroup while the majority (n = 61) were non-groupable. Throat sample is clearly the sampling method of choice, in 56 individuals where both throat and nasopharynx samples were taken, N. meningitidis was detected in both throat and nasopharynx in eight individuals, in 46 individuals N. meningitidis was only detected in the throat and in two individuals only in the nasopharynx. Carriage studies are important to provide knowledge of the current epidemiology and association between carrier isolates and disease-causing isolates in a given population. Therefore, planning for a carriage study in Sweden is in progress.

  • 540. Jahan, Mahabuba
    et al.
    Eriksson, Olof
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology, Radiology.
    Johnström, Peter
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Sundin, Anders
    Johansson, Lars
    Halldin, Christer
    Decreased defluorination by using the novel beta cell imaging agent [18F]FE-DTBZ-d4 in pigs examined by PETManuscript (preprint) (Other academic)
    Abstract [en]

    Introduction: Fluorine-18 DTBZ-analogues, which selectively targets the vesicular monoamine transporter 2 (VMAT2), have been extensively studied for in vivo quantification of beta cell mass by positron emission tomography (PET).  This study describes a novel deuterated radioligand [18F]FE-(+)-DTBZ-d4, aimed to increase the stability against in vivo defluorination previously observed for [18F]FE-(+)-DTBZ.

    Methods: [18F]FE-(+)-DTBZ-d4 was synthesized by alkylation of desmethyl -(+)-DTBZ precursor with deuterated  [18F]fluoroethyl bromide ([18F]FCD2CD2Br). Radioligand affinity and specificity to VMAT2 was assessed by an in vitro saturation homogenate binding assay using human endocrine and exocrine pancreatic tissues. In vivo PK/PD was studied in a porcine model by PET/CT. The rate of defluorination was quantified by compartmental modeling and contrasted against defluorination of the non-deuterated analogue.

    Results: [18F]FE-DTBZ-d4 was produced in good radiochemical yield (3.0-1.7 GBq) in 100 min. Radiochemical purity of the formulated product was > 98% for up to 5h. The in vitro Binding Potential (BP) for VMAT2 in islet tissue was 27.0±8.8. The BP was lower in exocrine tissue (1.7±1.0) in addition to a close to three-fold decrease in specificity. The rate of in vivo defluorination was decreased significantly (kdefluorination= 0.0016±0.0007) compared to the non-deuterated analogue (kdefluorination= 0.012±0.002), resulting in a more than six-fold increase in half-life stability.

    Conclusion: [18F]FE-(+)-DTBZ-d4 has favorable pharmacokinetic (PK) properties for VMAT2 imaging, in addition to gaining significantly increased stability against defluorination. The in vitro islet BP and specificity was lower compared to a non-deuterated analogue but the islet/exocrine BP ratio was unchanged, potentially allowing for improved target tissue discrimination.

  • 541. Jahan, Mahabuba
    et al.
    Eriksson, Olof
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Radiology.
    Johnström, Peter
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Sundin, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Radiology.
    Johansson, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Radiology.
    Halldin, Christer
    Decreased defluorination using the novel beta-cell imaging agent [18F]FE-DTBZ-d4 in pigs examined by PET2011In: EJNMMI research, ISSN 2191-219X, Vol. 1, no 1, p. 33-Article in journal (Refereed)
    Abstract [en]

    Background

    Fluorine-18 dihydrotetrabenazine [DTBZ] analogues, which selectively target the vesicular monoamine transporter 2 [VMAT2], have been extensively studied for in vivo quantification of beta cell mass by positron-emission tomography [PET]. This study describes a novel deuterated radioligand [18F]fluoroethyl [FE]-DTBZ-d4, aimed to increase the stability against in vivo defluorination previously observed for [18F]FE-DTBZ.

    Methods

    [18F]FE-DTBZ-d4 was synthesized by alkylation of 9-O-desmethyl-(+)-DTBZ precursor with deuterated [18F]FE bromide ([18F]FCD2CD2Br). Radioligand binding potential [BP] was assessed by an in vitro saturation homogenate binding assay using human endocrine and exocrine pancreatic tissues. In vivo pharmacokinetics and pharmacodynamics [PK/PD] was studied in a porcine model by PET/computed tomography, and the rate of defluorination was quantified by compartmental modeling.

    Results

    [18F]FE-DTBZ-d4 was produced in reproducible good radiochemical yield in 100 ± 20 min. Radiochemical purity of the formulated product was > 98% for up to 5 h with specific radioactivities that ranged from 192 to 529 GBq/μmol at the end of the synthesis. The in vitro BP for VMAT2 in the islet tissue was 27.0 ± 8.8, and for the exocrine tissue, 1.7 ± 1.0. The rate of in vivo defluorination was decreased significantly (kdefluorination = 0.0016 ± 0.0007 min-1) compared to the non-deuterated analogue (kdefluorination = 0.012 ± 0.002 min-1), resulting in a six fold increase in half-life stability.

    Conclusions

    [18F]FE-DTBZ-d4 has similar PK and PD properties for VMAT2 imaging as its non-deuterated analogue [18F]FE-DTBZ in addition to gaining significantly increased stability against defluorination. [18F]FE-DTBZ-d4 is a prime candidate for future preclinical and clinical studies on focal clusters of beta cells, such as in intramuscular islet grafts.

  • 542. Jahnmatz, Peter
    et al.
    Bengtsson, Theresa
    Zuber, Bartek
    Farnert, Anna
    Ahlborg, Niklas
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Mabtech AB, Sweden.
    An antigen-specific, four-color, B-cell FluoroSpot assay utilizing tagged antigens for detection2016In: JIM - Journal of Immunological Methods, ISSN 0022-1759, E-ISSN 1872-7905, Vol. 433, p. 23-30Article in journal (Refereed)
    Abstract [en]

    The FluoroSpot assay, a variant of ELISpot utilizing fluorescent detection, has so far been used primarily for assessment of T cells, where simultaneous detection of several cytokines has allowed a more qualitative analysis of functionally distinct T cells. The potential to measure multiple analytes also presents several advantages when analyzing B cells. Our aim was to develop a B-cell FluoroSpot assay adaptable to studies of a variety of antigens. The assay utilizes anti-IgG antibodies immobilized in 96-well filter membrane plates. During cell culture, IgG antibodies secreted by antibody-secreting cells (ASCs) are captured in the vicinity of each of these cells and the specificity of single ASCs is defined using antigens for detection. The antigens were labeled with biotin or peptide tags enabling secondary detection with fluorophore-conjugated streptavidin or tag-specific antibodies. The assay, utilizing up to four different tag systems and fluorophores simultaneously, was evaluated using hybridomas and immunized splenocytes as ASCs. Assay variants were developed that could: i) identify multiple ASCs with different antigen specificities; ii) detect ASCs showing cross-reactivity with different but related antigens; and iii) define the antigen-specificity and, by including anti-IgG subclass detection reagents, simultaneously determine the IgG subclass of antibodies secreted by ASCs. As demonstrated here, the B-cell FluoroSpot assay using tag-based detection systems provides a versatile and powerful tool to investigate antibody responses by individual cells that can be readily adapted to studies of a variety of antigen-specific ASCs.

  • 543.
    Jahns, Anika C.
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Lundskog, Bertil
    Umeå University, Faculty of Medicine.
    Berg, Johanna
    Umeå University, Faculty of Medicine.
    Jonsson, Rebecca
    Umeå University, Faculty of Medicine.
    McDowell, Andrew
    Patrick, Sheila
    Golovleva, Irina
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Medical and Clinical Genetics.
    Palmer, Ruth H.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Alexeyev, Oleg A.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Microbiology of folliculitis: a histological study of 39 cases2014In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 122, no 1, p. 25-32Article in journal (Refereed)
    Abstract [en]

    Folliculitis is a common inflammatory skin syndrome. Several microbial organisms have been put forward as causative agents, but few studies visualized microbes directly in inflamed hair follicles. This retrospective study investigated bacterial and fungal colonization of inflamed hair follicles in patients with clinically diagnosed non-infectious folliculitis. Skin biopsies from 39 folliculitis patients and 27 controls were screened by fluorescence in situ hybridization (FISH) using broad-range bacterial and fungal probes and by immunofluorescence microscopy using a monoclonal antibody towards Gram-positive bacteria. Specific monoclonal and polyclonal antibodies towards Staphylococcus spp. and Propionibacterium acnes were applied for further species identification. Inflamed follicles were associated with bacterial colonization in 10 samples (26%) and fungal colonization in three samples (8%). Staphylococcus spp. were observed in inflamed follicles in seven samples (18%). Two samples were positive for P. acnes, which were identified as either type II or type IB/type III. Both Staphylococcus spp. and P. acnes were seen in macrocolonies/biofilm structures. In conclusion, one-third of patients with clinically diagnosed, non-infectious folliculitis exhibited microbial colonization with predominance of Staphylococcus spp.

  • 544. James, Hayley R
    et al.
    Perzanowski, Matthew S.
    Rönmark, Eva
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine.
    Hedman, Linnéa
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine.
    Bjerg, Anders
    Schuyler, Alexander J
    Workman, Lisa J
    Lundback, Bo
    Platts-Mills, Thomas A E
    IgE Antibodies to Mammalian Allergens Are a Major Risk Factor for Prevalence, Severity, and Persistence of Asthma in Northern Sweden2015In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 135, no 2, p. AB22-AB22Article in journal (Other academic)
    Abstract [en]

    IgE to mammalian allergens can contribute significantly to asthma risk. Studying the details of the relationship between animal sensitization and asthma is simpler in an environment where mite, fungal, and cockroach allergens make little or no contribution to asthma risk. Methods: Quantitative assays for IgE to eight allergens were carried out on 963 sera from 19-year-olds in a population-based cohort in northern Sweden, and associations with questionnaire data from ages 7, 12, and 19 on asthma symptoms, diagnosis, and treatment were tested. Results: Overall, 79 (53%) of the students with a physician diagnosis of asthma were positive to one or more of the mammalian allergens (cat, dog, or horse danders) tested. Of the allergens assessed, only mammalian allergens, birch, and timothy grass pollen showed a significant relationship with asthma diagnosis. Multivariate analysis showed that high titer (>17.5 IU/ml) IgE to any mammalian allergen had the strongest relationship with asthma at age 19 (odds ratio 5.1 [3.0-8.6]). Furthermore, IgE to mammalian allergens gave an odds ratio of 8.5 [4.9-15] for asthma that started before age 12 and was still present at age 19. Sensitization to Fel d 1 and Fel d 4 was strongly associated with asthma and significantly reduced in cat owners. Conclusions: Sensitization to cat and dog related allergens, and specifically to the components Fel d 1 and Fel d 4, is a major risk factor for the persistence and severity of asthma in an area where these are the only significant perennial allergens.

  • 545.
    Jarefors, Sara
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Immunology . Linköping University, Faculty of Health Sciences.
    Cytokine responses in human Lyme borreliosis: The role of T helper 1-like immunity and aspects of gender and co-exposure in relation to disease course2006Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Lyme borreliosis was first described some 30 years ago in the USA. Today, it is the most common vector borne disease in Europe and the USA. The disease can have multiple stages and symptoms can manifest from various parts of the body; joints, skin heart and nervous system. In Europe, neuroborreliosis is the most frequent late stage diagnosis. Although Lyme borreliosis is treatable with antibiotics and the causative spirochete has not been shown to be resistant to drugs, some patients do not recover completely. They have persistent symptoms and are diagnosed with chronic or persistent Lyme borreliosis. The mechanism behind the lingering symptoms is unclear but might be due to tissue damage caused by the immune system. The aim of this thesis was to study the immunological differences between patients with different outcome of Lyme borreliosis, i.e. chronic, subacute and asymptomatic, and various factors that might influence the course of the disease.

    The Borrelia-specific IFN-γ and IL-4 secretion was detected in blood and cerebrospinal fluid from patients with chronic and subacute neuroborreliosis during the course of the disease. Blood samples were also obtained from patients with erythema migrans (EM) and acrodermatitis chronicum atrophicans. An early increase of IFN-γ with a later switch to an IL-4 response was observed in patients with a subacute disease course whereas the IFN-γ secretion continued to be elevated in chronic patients.

    The Borrelia-specific Th1-response was further investigated in chronic, subacute and asymptomatic individuals by studying the expression of the Th1-marker IL-12Rβ2, on a protein and mRNA level. The cytokine secretion and Foxp3, a marker for regulatory T-cells, were also analyzed. Chronic patients had a lower IL-12Rβ2 expression on CD8+ T-cells and a lower number of Borrelia-specific IFN-γ secreting cells compared to asymptomatic individuals. Chronic patients also displayed a higher expression of Borrelia-specific Foxp3 than healthy controls.

    The conclusions for these tow studies were that a strong Th1-response early in the infection with a later switch to a Th2-response is beneficiary whereas a slow or weak Th1-response corresponds to a prolonged disease course.

    The influence of a previous infection with another pathogen, seen to suppress the immune response in animals, and the possible gender difference in immune response was also investigated. Patients with EM were screened for antibodies to Anaplasma phagocytophilum (Ap) as a sign of a previous exposure to these tick-borne bacteria. Blood lymphocytes from Ap seronegative, Ap seropositive and healthy controls were stimulated with Borrelia antigen and the secretion of IL-4, IL-5, IL-12, IL-13 and IFN-γ was detected by ELISPOT. Ap seropositive patients had a lower number of cells responding with IL-12 secretion compared to the other groups which might indicate an inhibited Th1-response.

    Reinfections with Lyme borreliosis was in a previous study, done by Bennet et al, found to be more frequent in postmenopausal women than in men. To investigate if there was an immunological explanation to the gender discrepancy, blood lymphocytes from individuals reinfected with Lyme borreliosis and individuals infected only once were stimulated with various antigens. The cytokine secretion was detected by ELISPOT, ELISA and Immulite. There were no differences between reinfected and single infected individuals. However, women, regardless of times infected, displayed a Th2-derived and anti-inflammatory spontaneous immune response compared to men.

    A previous infection with the bacteria Ap might possibly have a long term effect on the immune system and might be of disadvantage when mounting a Th1-response to a Borrelia infection. Also, the Th2-derived response displayed by postmenopausal women could indicate why more women than men get reinfected with Borrelia burgdorferi.

  • 546. Javaheri, Anahita
    et al.
    Kruse, Tobias
    Moonens, Kristof
    Mejias-Luque, Raquel
    Debraekeleer, Ayla
    Asche, Carmen I.
    Tegtmeyer, Nicole
    Kalali, Behnam
    Bach, Nina C.
    Sieber, Stephan A.
    Hill, Darryl J.
    Koeniger, Verena
    Hauck, Christof R.
    Moskalenko, Roman
    Haas, Rainer
    Busch, Dirk H.
    Klaile, Esther
    Slevogt, Hortense
    Schmidt, Alexej
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology. Medical Faculty, Institute of Anatomy, University Duisburg-Essen, 45122 Essen, Germany.
    Backert, Steffen
    Remaut, Han
    Singer, Bernhard B.
    Gerhard, Markus
    Helicobacter pylori adhesin HopQ engages in a virulence-enhancing interaction with human CEACAMs2017In: Nature Microbiology, E-ISSN 2058-5276, Vol. 2, no 1, article id 16189Article in journal (Refereed)
    Abstract [en]

    Helicobacter pylori specifically colonizes the human gastric epithelium and is the major causative agent for ulcer disease and gastric cancer development. Here, we identify members of the carcinoembryonic antigen-related cell adhesion molecule (CEACAM) family as receptors of H. pylori and show that HopQ is the surface-exposed adhesin that specifically binds human CEACAM1, CEACAM3, CEACAM5 and CEACAM6. HopQ-CEACAM binding is glycan-independent and targeted to the N-domain. H. pylori binding induces CEACAM1-mediated signalling, and the HopQ-CEACAM1 interaction enables translocation of the virulence factor CagA into host cells and enhances the release of pro-inflammatory mediators such as interleukin-8. Based on the crystal structure of HopQ, we found that a beta-hairpin insertion (HopQ-ID) in HopQ's extracellular 3+4 helix bundle domain is important for CEACAM binding. A peptide derived from this domain competitively inhibits HopQ-mediated activation of the Cag virulence pathway, as genetic or antibody-mediated abrogation of the HopQ function shows. Together, our data suggest the HopQ-CEACAM1 interaction to be a potentially promising novel therapeutic target to combat H. pylori-associated diseases.

  • 547.
    Jayaprakash, Kartheyaene
    et al.
    Örebro University, School of Medical Sciences.
    Demirel, Isak
    Örebro University, School of Medical Sciences.
    Gunaltay, Sezin
    Örebro University, School of Medical Sciences.
    Khalaf, Hazem
    Örebro University, School of Medical Sciences.
    Bengtsson, Torbjörn
    Örebro University, School of Medical Sciences.
    PKC, ERK/p38 MAP kinases and NF-B targeted signalling play a role in the expression and release of IL-1β  and CXCL8 in Porphyromonas gingivalis-infected THP1 cells2017In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 125, no 7, p. 623-633Article in journal (Refereed)
    Abstract [en]

    Porphyromonas gingivalis is a keystone pathogen in periodontitis and is gaining importance in cardiovascular pathogenesis. Protease-activated receptors (PARs), toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD) on monocytes recognize the structural components on P. gingivalis, inducing inflammatory intermediates. Here, we elucidate the modulation of PARs, TLRs, NODs, and the role of MAPK and NF-B in IL-1 and CXCL8 release. THP1 cells were stimulated with P. gingivalis wild-type W50 and its isogenic gingipain mutants: Rgp mutant E8 and Kgp mutant K1A. We observed modulation of PARs, TLRs, NOD, IL-1 and CXCL8 expression by P. gingivalis. Gingipains hydrolyse IL-1 and CXCL8, which is more evident for IL-1 accumulation at 24 h. Inhibition of PKC (protein kinase C), p38 and ERK (extracellular signal-regulated kinases) partially reduced P. gingivalis-induced IL-1 at 6 h, whereas PKC and ERK reduced CXCL8 at both 6 and 24 h. Following NF-B inhibition, P. gingivalis-induced IL-1 and CXCL8 were completely suppressed to basal levels. Overall, TLRs, PARs and NOD possibly act in synergy with PKC, MAPK ERK/p38 and NF-B in P. gingivalis-induced IL-1 and CXCL8 release from THP1 cells. These pro-inflammatory cytokines could affect leucocytes in circulation and exacerbate other vascular inflammatory conditions such as atherosclerosis.

  • 548.
    Jayaprakash, Kartheyaene
    et al.
    Department of Medical Sciences, Örebro University, Örebro, Sweden.
    Demirel, Isak
    Örebro University, School of Medical Sciences.
    Khalaf, Hazem
    Örebro University, School of Medical Sciences.
    Bengtsson, Torbjörn
    Örebro University, School of Medical Sciences.
    Porphyromonas gingivalis-induced inflammatory responses in THP1 cells are altered by native and modified low-density lipoproteins in a strain-dependent manner2018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 8, p. 667-677Article in journal (Refereed)
    Abstract [en]

    Strong epidemiological evidence supports an association between cardiovascular and periodontal disease and furthermore, the periodontopathogen Porphyromonas gingivalis has been identified in blood and from atheromatous plaques. Blood exposed to P.gingivalis shows an increased protein modification of low-density lipoprotein (LDL). In this study, we investigate the inflammatory responses of THP1 cells incubated with P.gingivalis and the effects of native or modified LDL on these responses. Reactive oxygen species (ROS) and IL-1 were observed in THP1 cells following infection with P.gingivalis ATCC33277 and W50. Caspase 1 activity was quantified in THP1 cells and correlated with IL-1 accumulation. Oxidized LDL (oxLDL) induced IL-1 release and CD36 expression on THP1 cells. Modified LDL co-stimulated with ATCC33277 exhibited regulatory effects on caspase 1 activity, IL-1 release and CD36 expression in THP1 cells, whereas W50 induced more modest responses in THP1 cells. In summary, we show that P.gingivalis is capable of inducing pro-inflammatory responses in THP1 cells, and native and modified LDL could alter these responses in a dose- and strain-dependent manner. Strain-dependent differences in THP1 cell responses could be due to the effect of P.gingivalis proteases, presence or absence of capsule and proteolytic transformation of native and modified LDL.

  • 549.
    Jin, Chuan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University.
    Improvement of adoptive T-cell therapy for Cancer2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Cancer immunotherapy has recently made remarkable clinical progress. Adoptive transfer of T-cells engineered with a chimeric antigen receptor (CAR) against CD19 has been successful in treatment of B-cell leukemia. Patient’s T-cells are isolated, activated, transduced with a vector encoding the CAR molecule and then expanded before being transferred back to the patient. However some obstacles restrict its success in solid tumors. This thesis explores different aspects to improve CAR T-cells therapy of cancer.

    Ex vivo expanded T-cells are usually sensitive to the harsh tumor microenvironment after reinfusion. We developed a novel expansion method for T-cells, named AEP, by using irradiated and preactivated allo-sensitized allogeneic lymphocytes (ASALs) and allogeneic mature dendritic cells (DCs). AEP-expanded T-cells exhibited better survival and cytotoxic efficacy under oxidative and immunosuppressive stress, compared to T-cells expanded with established procedures.

    Integrating retro/lentivirus (RV/LV) used for CAR expressions randomly integrate in the T-cell genome and has the potential risk of causing insertional mutagenesis. We developed a non-integrating lentiviral (NILV) vector containing a scaffold matrix attachment region (S/MAR) element (NILV-S/MAR) for T-cells transduction. NILV-S/MAR-engineered CAR T-cells display similar cytotoxicity to LV-engineered CAR T-cells with undetectable level of insertional event, which makes them safer than CAR T-cells used in the clinic today.

    CD19-CAR T-cells have so far been successful for B-cell leukemia but less successful for B-cell lymphomas, which present semi-solid structure with an immunosuppressive microenvironment. We have developed CAR T-cells armed with H. pylorineutrophil-activating protein (HP-NAP). HP-NAP is a major virulence factor and plays important role in T-helper type 1 (Th1) polarizing. NAP-CAR T-cells showed the ability to mature DCs, attract innate immune cells and increase secretion of Th1 cytokines and chemokines, which presumably leads to better CAR T-cell therapy for B-cell lymphoma.

    Allogeneic-DCs (alloDCs) were used to further alter tumor microenvironment. The premise relies on initiation of an allo-reactive immune response for cytokine and chemokines secretion, as well as stimulation of T-cell response by bringing in tumor-associated antigen. We demonstrated that alloDCs promote migration and activation of immune cells and prolong the survival of tumor-bearing mice by attracting T-cells to tumors and reverse the immune suppressive tumor microenvironment.

  • 550.
    Jin, Chuan
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Fotaki, Grammatiki
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Ramachandran, Mohanraj
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Nilsson, Berith
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Essand, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Yu, Di
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Safe engineering of CAR T cells for adoptive cell therapy of cancer using long-term episomal gene transfer2016In: EMBO Molecular Medicine, ISSN 1757-4676, E-ISSN 1757-4684, Vol. 8, no 7, p. 702-711Article in journal (Refereed)
    Abstract [en]

    Chimeric antigen receptor (CAR) T-cell therapy is a new successful treatment for refractory B-cell leukemia. Successful therapeutic outcome depends on long-term expression of CAR transgene in T cells, which is achieved by delivering transgene using integrating gamma retrovirus (RV) or lentivirus (LV). However, uncontrolled RV/LV integration in host cell genomes has the potential risk of causing insertional mutagenesis. Herein, we describe a novel episomal long-term cell engineering method using non-integrating lentiviral (NILV) vector containing a scaffold/matrix attachment region (S/MAR) element, for either expression of transgenes or silencing of target genes. The insertional events of this vector into the genome of host cells are below detection level. CD19 CAR T cells engineered with a NILV-S/MAR vector have similar levels of CAR expression as T cells engineered with an integrating LV vector, even after numerous rounds of cell division. NILV-S/MAR-engineered CD19 CAR T cells exhibited similar cytotoxic capacity upon CD19(+) target cell recognition as LV-engineered T cells and are as effective in controlling tumor growth in vivo We propose that NILV-S/MAR vectors are superior to current options as they enable long-term transgene expression without the risk of insertional mutagenesis and genotoxicity.

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