| 1. |
- Ahrens, Norbert, et al.
(författare)
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Mesenchymal stem cell content of human vertebral bone marrow
- 2004
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Ingår i: Transplantation. - 1534-6080. ; 78:6, s. 925-929
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Tidskriftsartikel (refereegranskat)abstract
- Mesenchymal stem cells (MSCs) are capable of down-regulating alloimmune responses and promoting the engraftment of hematopoietic stem cells. MSCs may therefore be suitable for improving donor-specific tolerance induction in solid-organ transplantation. Cells from cadaveric vertebral bone marrow (V-BM), aspirated iliac crest-BM, and peripheral blood progenitor cells were compared. Cells were characterized by flow cytometry and colony assays. MSCs generated from V-BM were assayed for differentiation capacity and immunomodulatory function. A median 5.7 x 10(8) nucleated cells (NCs) were recovered per vertebral body. The mesenchymal progenitor, colony-forming unit-fibroblast, frequency in V-BM (11.6/10(5) NC, range: 6.0-20.0) was considerably higher than in iliac crest-BM (1.4/10(5) NC, range: 0.4-2.6) and peripheral blood progenitor cells (not detectable). MSC generated from V-BM had the typical MSC phenotype (CD105(pos)CD73(pos)CD45(neg)CD34(neg)), displayed multilineage differentiation potential, and suppressed alloreactivity in mixed lymphocyte reactions. V-BM may be an excellent source for MSC cotransplantation approaches.
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| 2. |
- Brogden, Graham, et al.
(författare)
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Methods to study lipid alterations in neutrophils and the subsequent formation of neutrophil extracellular traps
- 2017
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Ingår i: Journal of Visualized Experiments. - : MyJove Corporation. - 1940-087X. ; 2017:121
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Tidskriftsartikel (refereegranskat)abstract
- Lipid analysis performed by high performance thin layer chromatography (HPTLC) is a relatively simple, cost-effective method of analyzing a broad range of lipids. The function of lipids (e.g., in host-pathogen interactions or host entry) has been reported to play a crucial role in cellular processes. Here, we show a method to determine lipid composition, with a focus on the cholesterol level of primary blood-derived neutrophils, by HPTLC in comparison to high performance liquid chromatography (HPLC). The aim was to investigate the role of lipid/cholesterol alterations in the formation of neutrophil extracellular traps (NETs). NET release is known as a host defense mechanism to prevent pathogens from spreading within the host. Therefore, blood-derived human neutrophils were treated with methyl-β-cyclodextrin (MβCD) to induce lipid alterations in the cells. Using HPTLC and HPLC, we have shown that MβCD treatment of the cells leads to lipid alterations associated with a significant reduction in the cholesterol content of the cell. At the same time, MβCD treatment of the neutrophils led to the formation of NETs, as shown by immunofluorescence microscopy. In summary, here we present a detailed method to study lipid alterations in neutrophils and the formation of NETs.
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| 3. |
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| 4. |
- Frick, Inga Maria, et al.
(författare)
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Streptococcal inhibitor of complement (SIC) modulates fibrinolysis and enhances bacterial survival within fibrin clots
- 2018
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Ingår i: Journal of Biological Chemistry. - 0021-9258. ; 293:35, s. 13578-13591
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Tidskriftsartikel (refereegranskat)abstract
- Some strains of the bacterial pathogen Streptococcus pyogenes secrete protein SIC (streptococcal inhibitor of complement), including strains of the clinically relevant M1 serotype. SIC neutralizes the effect of a number of antimicrobial proteins/peptides and interferes with the function of the host complement system. Previous studies have shown that some S. pyogenes proteins bind and modulate coagulation and fibrinolysis factors, raising the possibility that SIC also may interfere with the activity of these factors. Here we show that SIC interacts with both human thrombin and plasminogen, key components of coagulation and fibrinolysis. We found that during clot formation, SIC binds fibrin through its central region and that SIC inhibits fibrinolysis by interacting with plasminogen. Flow cytometry results indicated that SIC and plasminogen bind simultaneously to S. pyogenes bacteria, and fluorescence microscopy revealed co-localization of the two proteins at the bacterial surface. As a consequence, SIC-expressing bacteria entrapped in clots inhibit fibrinolysis, leading to delayed bacterial escape from the clots as compared with mutant bacteria lacking SIC. Moreover, within the clots SIC-expressing bacteria were protected against killing. In an animal model of subcutaneous infection, SIC-expressing bacteria exhibited a delayed systemic spread. These results demonstrate that the bacterial protein SIC interferes with coagulation and fibrinolysis and thereby enhances bacterial survival, a finding that has significant implications for S. pyogenes virulence.
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| 5. |
- Grasso, Simona, et al.
(författare)
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Interaction of factor VII activating protease (FSAP) with neutrophil extracellular traps (NETs)
- 2018
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Ingår i: Thrombosis Research. - : Elsevier BV. - 0049-3848. ; 161, s. 36-42
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Tidskriftsartikel (refereegranskat)abstract
- The circulating zymogen form of Factor VII activating protease (FSAP) can be activated by histones and nucleosomes in vivo. These cell-death-associated nuclear factors are also actively extruded into the extracellular space by neutrophils through a process called neutrophil extracellular trap (NET) formation (NETosis). NETs are thought to be involved in host defense, inflammation as well as thrombosis. We have investigated the bidirectional interactions of FSAP and NETs. Phorbol ester-mediated NET formation was marginally stimulated by FSAP. Plasma-derived FSAP as well as exogenous FSAP bound to NETs. There was co-localization of FSAP and NETs in coronary thrombi from patients with acute myocardial infarction. Contrary to our expectations no activation of pro-FSAP by NETs was evident. However, after disintegration of NETs with DNase, a robust activation of pro-FSAP, due to release of histones from nucleosomes, was detected. The released histones were in turn degraded by FSAP. Histone cytotoxicity towards endothelial cells was neutralized by FSAP more potently than by activated protein C (APC). One more consequence of histone degradation was a decrease in nucleosome release from apoptotic neutrophils. Taken together, NETs bind to FSAP, but do not activate pro-FSAP unless histones are released from NETs by DNAse. This activation of FSAP is likely to be important in diminishing the cytotoxic effect of histones, thus limiting the damaging effect of NETosis.
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| 6. |
- Mohanty, Tirthankar, et al.
(författare)
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Neutrophil extracellular traps in the central nervous system hinder bacterial clearance during pneumococcal meningitis
- 2019
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 10:1, s. 1667-1667
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophils are crucial mediators of host defense that are recruited to the central nervous system (CNS) in large numbers during acute bacterial meningitis caused by Streptococcus pneumoniae. Neutrophils release neutrophil extracellular traps (NETs) during infections to trap and kill bacteria. Intact NETs are fibrous structures composed of decondensed DNA and neutrophil-derived antimicrobial proteins. Here we show NETs in the cerebrospinal fluid (CSF) of patients with pneumococcal meningitis, and their absence in other forms of meningitis with neutrophil influx into the CSF caused by viruses, Borrelia and subarachnoid hemorrhage. In a rat model of meningitis, a clinical strain of pneumococci induced NET formation in the CSF. Disrupting NETs using DNase I significantly reduces bacterial load, demonstrating that NETs contribute to pneumococcal meningitis pathogenesis in vivo. We conclude that NETs in the CNS reduce bacterial clearance and degrading NETs using DNase I may have significant therapeutic implications.
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| 7. |
- Neumann, Ariane, et al.
(författare)
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Extracellular traps : An ancient weapon of multiple kingdoms
- 2020
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Ingår i: Biology. - : MDPI AG. - 2079-7737. ; 9:2
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Forskningsöversikt (refereegranskat)abstract
- The discovery, in 2004, of extracellular traps released by neutrophils has extended our understanding of the mode of action of various innate immune cells. This fascinating discovery demonstrated the extracellular trapping and killing of various pathogens by neutrophils. During the last decade, evidence has accumulated showing that extracellular traps play a crucial role in the defence mechanisms of various cell types present in vertebrates, invertebrates, and plants. The aim of this review is to summarise the relevant literature on the evolutionary history of extracellular traps used as a weapon in various kingdoms of life.
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| 8. |
- Neumann, Ariane, et al.
(författare)
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Finegoldia magna, an Anaerobic Gram-Positive Bacterium of the Normal Human Microbiota, Induces Inflammation by Activating Neutrophils
- 2020
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Ingår i: Frontiers in Microbiology. - : Frontiers Media SA. - 1664-302X. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- The Gram-positive anaerobic commensal Finegoldia magna colonizes the skin and other non-sterile body surfaces, and is an important opportunistic pathogen. Here we analyzed the effect of F. magna on human primary neutrophils. F. magna strains ALB8 (expressing protein FAF), 312 (expressing protein L) and 505 (naturally lacking both protein FAF and L) as well as their associated proteins activate neutrophils to release reactive oxygen species, an indication for neutrophil oxidative burst. Co-incubation of neutrophils with the bacteria leads to a strong increase of CD66b surface expression, another indicator for neutrophil activation. Furthermore, all tested stimuli triggered the release of NETs from the activated neutrophils, pointing to a host defense mechanism in response to the tested stimuli. This phenotype is dependent on actin rearrangement, NADPH oxidases and the ERK1/2 pathway. Proteins FAF and L also induced the secretion of several pro-inflammatory neutrophil proteins; HBP, IL-8 and INFγ. This study shows for the first time a direct interaction of F. magna with human neutrophils and suggests that the activation of neutrophils plays a role in F. magna pathogenesis.
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| 9. |
- Neumann, Ariane, et al.
(författare)
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Immunoregulation of Neutrophil Extracellular Trap Formation by Endothelial-Derived p33 (gC1q Receptor)
- 2018
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Ingår i: Journal of Innate Immunity. - : S. Karger AG. - 1662-811X .- 1662-8128. ; 10:1, s. 30-43
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Tidskriftsartikel (refereegranskat)abstract
- The formation of neutrophil extracellular traps (NETs) is a host defence mechanism, known to facilitate the entrapment and growth inhibition of many bacterial pathogens. It has been implicated that the translocation of myeloperoxidase (MPO) from neutrophilic granules to the nucleus is crucial to this process. Under disease conditions, however, excessive NET formation can trigger self-destructive complications by releasing pathologic levels of danger-associated molecular pattern molecules (DAMPs). To counteract such devastating immune reactions, the host has to rely on precautions that help circumvent these deleterious effects. Though the induction of DAMP responses has been intensively studied, the mechanisms that are used by the host to down-regulate them are still not understood. In this study, we show that p33 is an endothelial-derived protein that has the ability to annul NET formation. We found that the expression of human p33 is up-regulated in endothelial cells upon infections with Streptococcus pyogenes bacteria. Using tissue biopsies from a patient with streptococcal necrotising fasciitis, we monitored co-localisation of p33 with MPO. Further in vitro studies revealed that p33 is able to block the formation of DAMP-induced NET formation by inhibiting the enzymatic activity of MPO. Additionally, mice challenged with S. pyogenes bacteria demonstrated diminished MPO activity when treated with p33. Together, our results demonstrate that host-derived p33 has an important immunomodulating function that helps to counterbalance an overwhelming DAMP response.
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| 10. |
- Neumann, Ariane, et al.
(författare)
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Novel Role of the Antimicrobial Peptide LL-37 in the Protection of Neutrophil Extracellular Traps against Degradation by Bacterial Nucleases
- 2014
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Ingår i: Journal of Innate Immunity. - : S. Karger AG. - 1662-811X .- 1662-8128. ; 6:6, s. 860-868
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil extracellular traps (NETs) have been described as a fundamental innate immune defence mechanism. They consist of a nuclear DNA backbone associated with different antimicrobial peptides (AMPs) which are able to engulf and kill pathogens. The AMP LL-37, a member of the cathelicidin family, is highly present in NETs. However, the function of LL-37 within NETs is still unknown because it loses its antimicrobial activity when bound to DNA in the NETs. Using immunofluorescence microscopy, we demonstrate that NETs treated with LL-37 are distinctly more resistant to S. aureus nuclease degradation than nontreated NETs. Biochemical assays utilising a random LL-37-fragment library indicated that the blocking effect of LL-37 on nuclease activity is based on the cationic character of the AMP, which facilitates the binding to neutrophil DNA, thus protecting it from degradation by the nuclease. In good correlation to these data, the cationic AMPs human beta defensin-3 and human neutrophil peptide-1 showed similar protection of neutrophil-derived DNA against nuclease degradation. In conclusion, this study demonstrates a novel role of AMPs in host immune defence: beside its direct antimicrobial activity against various pathogens, cationic AMPs can stabilise neutrophil-derived DNA or NETs against bacterial nuclease degradation. (C) 2014 S. Karger AG, Basel
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