| 1. |
- Serrander, Lena, et al.
(författare)
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Particles binding β2-integrins mediate intracellular production of oxidative metabolites in human neutrophils independently of phagocytosis
- 1999
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Ingår i: Biochimica et Biophysica Acta. Molecular Cell Research. - 0167-4889 .- 1879-2596. ; 1452:2, s. 133-144
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Tidskriftsartikel (refereegranskat)abstract
- Complement-opsonised particles are readily ingested by human neutrophils through a complement receptor-mediated process leading to phagolysosome fusion and production of oxidative metabolites. To investigate the complement receptor 3 (CR3)-associated signal system involved, cells were challenged with protein A-positive, heat-killed Staphylococcus aureus to which antibodies with specificity for the subunits of the β2-integrins, i.e. anti-CD11b (the α subunit of CR3) and anti-CD18 (the β subunit of CR3), were bound through their Fc moiety. Despite not being ingested by the neutrophils, the surface associated anti-CD18- and anti-CD11b-coated particles were able to activate the neutrophil NADPH-oxidase. Also anti-CD11a- (the α subunit of LFA-1) and to a lesser extent anti-CD11c- (the α subunit of CR4) coated particles were able to trigger the NADPH-oxidase. The NADPH-oxidase was activated without extracellular release of reactive oxygen species. The activity was inhibited by cytochalasin B, suggesting a necessary role for the cytoskeleton in the signalling pathway that activates the oxidase. We show that particle-mediated cross-linking of β2-integrins on the neutrophil surface initiates a signalling cascade, involving cytoskeletal rearrangements, leading to an activation of the NADPH-oxidase without phagosome formation or extracellular release of reactive oxygen species.
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| 2. |
- von Beek, Christopher, et al.
(författare)
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A two-step activation mechanism enables mast cells to differentiate their response between extracellular and invasive enterobacterial infection
- 2024
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Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- Mast cells localize to mucosal tissues and contribute to innate immune defense against infection. How mast cells sense, differentiate between, and respond to bacterial pathogens remains a topic of ongoing debate. Using the prototype enteropathogen Salmonella Typhimurium (S.Tm) and other related enterobacteria, here we show that mast cells can regulate their cytokine secretion response to distinguish between extracellular and invasive bacterial infection. Tissue-invasive S.Tm and mast cells colocalize in the mouse gut during acute Salmonella infection. Toll-like Receptor 4 (TLR4) sensing of extracellular S.Tm, or pure lipopolysaccharide, causes a modest induction of cytokine transcripts and proteins, including IL-6, IL-13, and TNF. By contrast, type-III-secretion-system-1 (TTSS-1)-dependent S.Tm invasion of both mouse and human mast cells triggers rapid and potent inflammatory gene expression and >100-fold elevated cytokine secretion. The S.Tm TTSS-1 effectors SopB, SopE, and SopE2 here elicit a second activation signal, including Akt phosphorylation downstream of effector translocation, which combines with TLR activation to drive the full-blown mast cell response. Supernatants from S.Tm-infected mast cells boost macrophage survival and maturation from bone-marrow progenitors. Taken together, this study shows that mast cells can differentiate between extracellular and host-cell invasive enterobacteria via a two-step activation mechanism and tune their inflammatory output accordingly.
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| 3. |
- von Beek, Christopher, et al.
(författare)
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A two-step activation mechanism enables mast cells to differentiate their response between extracellular and invasive enterobacterial infection
- 2024
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Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- Mast cells localize to mucosal tissues and contribute to innate immune defense against infection. How mast cells sense, differentiate between, and respond to bacterial pathogens remains a topic of ongoing debate. Using the prototype enteropathogen Salmonella Typhimurium (S.Tm) and other related enterobacteria, here we show that mast cells can regulate their cytokine secretion response to distinguish between extracellular and invasive bacterial infection. Tissue-invasive S.Tm and mast cells colocalize in the mouse gut during acute Salmonella infection. Toll-like Receptor 4 (TLR4) sensing of extracellular S.Tm, or pure lipopolysaccharide, causes a modest induction of cytokine transcripts and proteins, including IL-6, IL-13, and TNF. By contrast, type-III-secretion-system-1 (TTSS-1)-dependent S.Tm invasion of both mouse and human mast cells triggers rapid and potent inflammatory gene expression and >100-fold elevated cytokine secretion. The S.Tm TTSS-1 effectors SopB, SopE, and SopE2 here elicit a second activation signal, including Akt phosphorylation downstream of effector translocation, which combines with TLR activation to drive the full-blown mast cell response. Supernatants from S.Tm-infected mast cells boost macrophage survival and maturation from bone-marrow progenitors. Taken together, this study shows that mast cells can differentiate between extracellular and host-cell invasive enterobacteria via a two-step activation mechanism and tune their inflammatory output accordingly.
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| 4. |
- Weineisen, Maria, et al.
(författare)
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Streptococcal M5 protein prevents neutrophil phagocytosis by interfering with CD11b/CD18 receptor-mediated association and signaling.
- 2004
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Ingår i: J Immunol. - 0022-1767. ; 172:6, s. 3798-3807
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Tidskriftsartikel (refereegranskat)abstract
- Group A streptococci (GAS) are common human pathogens that express major surface-associated virulence factors designated M proteins. In this study, we explored directly the cellular mechanisms behind their supposed ability to prevent phagocytosis. Isolated human neutrophils killed an M-negative GAS mutant (DeltaM5), but not the wild-type parent strain (M5). After 3 h, 3-4 times as many DeltaM5 as M5 bacteria were associated with the neutrophils, and more DeltaM5 than M5 bacteria were ingested. However, there was no statistically significant difference between DeltaM5 and M5 bacteria in regard to the percentage of the neutrophil-associated bacteria that were ingested, indicating that M5 protein prevents an adhesion receptor-dependent association with neutrophils and not the phagocytic machinery per se. Different Abs against CD11b/CD18 (CR3) blocked adhesion and killing of DeltaM5 bacteria, whereas the blocking of two other complement receptors, CD11c/CD18 (CR4) and CD35 (CR1), did not. The CD11b/CD18-mediated killing of DeltaM5 bacteria resulted in protein tyrosine phosphorylations and Cdc42 activation. Furthermore, inhibition of CD11b/CD18 receptor engagement or tyrosine kinase activity blocked the DeltaM5-induced activation of Cdc42 as well as the killing of these bacteria. We conclude that M5 protein interferes with the CD11b/CD18-dependent association between GAS and neutrophils, and thereby blocks subsequent ingestion of the bacteria.
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| 5. |
- Akopyan, Karen, et al.
(författare)
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Translocation of surface-localized effectors in type III secretion
- 2011
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 108:4, s. 1639-1644
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Tidskriftsartikel (refereegranskat)abstract
- Pathogenic Yersinia species suppress the host immune response by using a plasmid-encoded type III secretion system (T3SS) to translocate virulence proteins into the cytosol of the target cells. T3SS-dependent protein translocation is believed to occur in one step from the bacterial cytosol to the target-cell cytoplasm through a conduit created by the T3SS upon target cell contact. Here, we report that T3SS substrates on the surface of Yersinia pseudotuberculosis are translocated into target cells. Upon host cell contact, purified YopH coated on Y. pseudotuberculosis was specifically and rapidly translocated across the target-cell membrane, which led to a physiological response in the infected cell. In addition, translocation of externally added YopH required a functional T3SS and a specific translocation domain in the effector protein. Efficient, T3SS-dependent translocation of purified YopH added in vitro was also observed when using coated Salmonella typhimurium strains, which implies that T3SS-mediated translocation of extracellular effector proteins is conserved among T3SS-dependent pathogens. Our results demonstrate that polarized T3SS-dependent translocation of proteins can be achieved through an intermediate extracellular step that can be reconstituted in vitro. These results indicate that translocation can occur by a different mechanism from the assumed single-step conduit model.
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| 6. |
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| 7. |
- Andersson, Kerstin, et al.
(författare)
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Yersinia pseudotuberculosis-induced calcium signaling in neutrophils is blocked by the virulence effector YopH
- 1999
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Ingår i: Infection and Immunity. - 0019-9567 .- 1098-5522. ; 67:5, s. 2567-2574
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Tidskriftsartikel (refereegranskat)abstract
- Pathogenic species of the genus Yersinia evade the bactericidal functions of phagocytes. This evasion is mediated through their virulence effectors, Yops, which act within target cells. In this study we investigated the effect of Yersinia pseudotuberculosis on Ca 2+ signaling in polymorphonuclear neutrophils. The intracellular free calcium concentration in single adherent human neutrophils was monitored during bacterial infection and, in parallel, the encounter between the bacteria and cells was observed. When a plasmid-cured strain was used for infection, adherence of a single bacterium to the cellular surface induced a β 1 integrin-dependent transient increase in the intracellular concentration of free calcium. This was, however, not seen with Yop-expressing wild-type bacteria, which adhered to the cell surface without generating any Ca 2+ signal. Importantly, the overall Ca 2+ homeostasis was not affected by the wild-type strain; the Ca 2+ signal mediated by the G-protein-coupled formyl-methionyl-leucyl- phenylalanine receptor was still functioning. Hence, the blocking effect was restricted to certain receptors and their signaling pathways. The use of different Yop mutant strains revealed that the protein tyrosine phosphatase YopH was responsible for the inhibition. This virulence determinant has previously been implicated in very rapid Yersinia-mediated effects on target cells as the key effector in the blockage of phagocytic uptake. The present finding, that Y. pseudotuberculosis, via YopH, specifically inhibits a self- induced immediate-early Ca 2+ signal in neutrophils, offers more-detailed information concerning the effectiveness of this virulence effector and implies an effect on Ca 2+-dependent, downstream signals.
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| 8. |
- Antti, Henrik, et al.
(författare)
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Metabolic profiling for detection of staphylococcus aureus infection and antibiotic resistance
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:2
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Tidskriftsartikel (refereegranskat)abstract
- Due to slow diagnostics, physicians must optimize antibiotic therapies based on clinical evaluation of patients without specific information on causative bacteria. We have investigated metabolomic analysis of blood for the detection of acute bacterial infection and early differentiation between ineffective and effective antibiotic treatment. A vital and timely therapeutic difficulty was thereby addressed: the ability to rapidly detect treatment failures because of antibiotic-resistant bacteria. Methicillin-resistant (MRSA) and methicillin-sensitive (MSSA) were used and for infecting mice, while natural MSSA infection was studied in humans. Samples of bacterial growth media, the blood of infected mice and of humans were analyzed with combined Gas Chromatography/Mass Spectrometry. Multivariate data analysis was used to reveal the metabolic profiles of infection and the responses to different antibiotic treatments. experiments resulted in the detection of 256 putative metabolites and mice infection experiments resulted in the detection of 474 putative metabolites. Importantly, ineffective and effective antibiotic treatments were differentiated already two hours after treatment start in both experimental systems. That is, the ineffective treatment of MRSA using cloxacillin and untreated controls produced one metabolic profile while all effective treatment combinations using cloxacillin or vancomycin for MSSA or MRSA produced another profile. For further evaluation of the concept, blood samples of humans admitted to intensive care with severe sepsis were analyzed. One hundred thirty-three putative metabolites differentiated severe MSSA sepsis (n = 6) from severe sepsis (n = 10) and identified treatment responses over time. Combined analysis of human, , and mice samples identified 25 metabolites indicative of effective treatment of sepsis. Taken together, this study provides a proof of concept of the utility of analyzing metabolite patterns in blood for early differentiation between ineffective and effective antibiotic treatment in acute infections.
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| 9. |
- Avican, Kemal, 1980-
(författare)
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Persistent infection by Yersinia pseudotuberculosis
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Enteropathogenic Yersinia species can infect many mammalian organs such as the small intestine, cecum, Peyer’s patches, liver, spleen, and lung and cause diseases that resemble a typhoid-like syndrome, as seen for other enteropathogens. We found that sublethal infection doses of Y. pseudotuberculosis gave rise to asymptomatic persistent infection in mice and identified the cecal lymphoid follicles as the primary site for colonization during persistence. Persistent Y. pseudotuberculosis is localized in the dome area, often in inflammatory lesions, as foci or as single cells, and also in neutrophil exudates in the cecal lumen. This new mouse model for bacterial persistence in cecum has potential as an investigative tool for deeper understanding of bacterial adaptation and host immune defense mechanisms during persistent infection. Here, we investigated the nature of the persistent infection established by Y. pseudotuberculosis in mouse cecal tissue using in vivo RNA-seq of bacteria during early and persistent stages of infection. Comparative analysis of the bacterial transcriptomes revealed that Y. pseudotuberculosis undergoes transcriptional reprogramming with drastic down-regulation of T3SS virulence genes during persistence in the cecum. At the persistent stage, the expression pattern in many respects resembles the pattern seen in vitro at 26°C. Genes that are up-regulated during persistence are genes involved in anaerobiosis, chemotaxis, and protection against oxidative and acidic stress, which indicates the influence of different environmental cues. We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence. Furthermore, we show that ArcA, Fnr, FrdA, WrbA, RovA, and RfaH play critical roles in persistence. An extended investigation of the transcriptional regulator rfaH employing mouse infection studies, phenotypic characterizations, and RNA-seq transcriptomics analyses indicated that this gene product contributes to establishment of infection and confirmed that it regulates O-antigen biosynthesis genes in Y. pseudotuberculosis. The RNA-seq results also suggest that rfaH has a relatively global effect. Furthermore, we also found that the dynamics of the cecal tissue organization and microbial composition shows changes during different stages of the infection. Taken together, based on our findings, we speculate that this enteropathogen initiates infection by using its virulence factors in meeting the innate immune response in the cecal tissue. Later on, these factors lead to dysbiosis in the local microbiota and altered tissue organization. At later stages of the infection, the pathogen adapts to the environment in the cecum by reprogramming its transcriptome from a highly virulent mode to a more environmentally adaptable mode for survival and shedding. The in vivo transcriptomic analyses for essential genes during infections present strong candidates for novel targets for antimicrobials.
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| 10. |
- Avican, Kemal, et al.
(författare)
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Reprogramming of Yersinia from Virulent to Persistent Mode Revealed by Complex In Vivo RNA-seq Analysis
- 2015
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Ingår i: PLoS Pathogens. - : Public Library of Science (PLoS). - 1553-7366 .- 1553-7374. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- We recently found that Yersinia pseudotuberculosis can be used as a model of persistent bacterial infections. We performed in vivo RNA-seq of bacteria in small cecal tissue biopsies at early and persistent stages of infection to determine strategies associated with persistence. Comprehensive analysis of mixed RNA populations from infected tissues revealed that Y. pseudotuberculosis undergoes transcriptional reprogramming with drastic down-regulation of T3SS virulence genes during persistence when the pathogen resides within the cecum. At the persistent stage, the expression pattern in many respects resembles the pattern seen in vitro at 26oC, with for example, up-regulation of flagellar genes and invA. These findings are expected to have impact on future rationales to identify suitable bacterial targets for new antibiotics. Other genes that are up-regulated during persistence are genes involved in anaerobiosis, chemotaxis, and protection against oxidative and acidic stress, which indicates the influence of different environmental cues. We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence. Furthermore, arcA, fnr, frdA, and wrbA play critical roles in persistence. Our findings suggest a model for the life cycle of this enteropathogen with reprogramming from a virulent to an adapted phenotype capable of persisting and spreading by fecal shedding.
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