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Inflammasomes: defense guardians in host-microbe defence
Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The inflammasomes are emerging as key regulators of the innate immune response as they response to cellular infection, tissue damage and/or stress. Activated inflammasomes provide a signalling platform for caspase-1 activation which subsequently processes the maturation of interleukin (IL)-1β and IL-18, and in addition promotes cell death, named pyroptosis. These effector mechanisms of the inflammasome constitute an important part of the innate immune response in host-defence.

The aims of this thesis were to elucidate if the inflammasome is activated by specific pathogens, and if genetic variations lead to susceptibility to severe inflammatory diseases, such as blood stream infections. We found that the commensal pathogens Staphylococcus aureus and Propionibacterium acnes induce caspase-1 activation, the latter being a stronger activator. The data also propose that it is rather the bacterial ligands than the secreted toxins of P. acnes that induce inflammasome activation. Even though the S. aureus PVL-toxin induces a caspase-1 activation, involvement of other virulence factors are of importance. Upon S. aureus and P. acnes stimulation, inter-individual variations were found, implying that the characteristics of the host innate immune system, rather than the properties of the bacteria, are decisive for the inflammatory response. Indeed, healthy individuals with the combined polymorphisms of the NLRP3 inflammasome (C10X/Q705K) have higher levels of spontaneous IL-1β, and most intriguingly C10X polymorphism showed a strong correlation with patients with bacteremia.

In conclusion, these studies suggest that different pathogens activate the inflammasomes to various degrees. The genetics of the host innate immune system, rather than the properties of the bacteria, define the inflammatory response against these bacteria, and specific genetic variations such as C10X polymorphism can increase the susceptibility for disease development.

Place, publisher, year, edition, pages
Örebro: Örebro universitet , 2013. , p. 76
Series
Örebro Studies in Medicine, ISSN 1652-4063 ; 88
Keywords [en]
Innate immunity, inflammasome, leukocytes, cytokines, Staphylococcus aureus, Propionibacterium acnes, inflammation, infection, gene variants, polymorphism.
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
URN: urn:nbn:se:oru:diva-28523ISBN: 978-91-7668-923-3 (print)OAI: oai:DiVA.org:oru-28523DiVA, id: diva2:613890
Public defence
2013-06-05, Wilandersalen, M-huset, Universitetetssjukhuset Örebro, Grev Rosen gatan 18, Örebro, 09:00 (Swedish)
Opponent
Supervisors
Available from: 2013-04-02 Created: 2013-04-02 Last updated: 2017-10-17Bibliographically approved
List of papers
1. Two adult siblings with atypical Cryopyrin-associated periodic syndrome due to a novel M299V mutation in NLRP3
Open this publication in new window or tab >>Two adult siblings with atypical Cryopyrin-associated periodic syndrome due to a novel M299V mutation in NLRP3
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2010 (English)In: Arthritis and Rheumatism, ISSN 0004-3591, E-ISSN 1529-0131, Vol. 62, no 7, p. 2138-2143Article in journal (Refereed) Published
Abstract [en]

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

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

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

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

National Category
Medical and Health Sciences
Research subject
Medicine
Identifiers
urn:nbn:se:oru:diva-12863 (URN)10.1002/art.27489 (DOI)000282758500034 ()
Note

ARTHRITIS AND RHEUMATISM ended 2013

Available from: 2011-01-10 Created: 2011-01-03 Last updated: 2017-12-11Bibliographically approved
2. Propionibacterium acnes activates caspase-1 in human neutrophils
Open this publication in new window or tab >>Propionibacterium acnes activates caspase-1 in human neutrophils
2013 (English)In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 121, no 7, p. 652-63Article in journal (Refereed) Published
Abstract [en]

Propionibacterium acnes is a Gram-positive, slow-growing, anaerobic bacillus, predominantly found as a commensal on the skin and mucous membranes of adults. It is, however, also considered an opportunistic pathogen; mostly associated with acne vulgaris, but rarely also with severe infections such as infective endocarditis, prosthetic joint infections, and deep sternal wound infections following cardiothoracic surgery. In addition, P. acnes has recently been found in high frequency in prostate tissue from patients with prostatitis and prostate cancer. The NOD-like receptors (NLR) act as intracellular sensors of microbial components, and a number of various bacteria have been found to induce assembling and activation of NLR-inflammasomes; leading to a pro-inflammatory response. The inflammasome-mediated formation of the pro-inflammatory cytokines interleukin-1β (IL-1β) and IL-18 involves the auto-proteolytic maturation of caspase-1. This study investigated if P. acnes activates inflammasomes. Propionibacterium acnes isolates (n = 29) with diverse origin were used as stimuli for peripheral leukocytes obtained from blood donors (BDs). The activity of inflammasomes was determined by measuring caspase-1 by flow cytometry and cytokine production by ELISA. A significant amount of caspase-1 was found in neutrophils upon P. acnes stimulation, whereas only a modest activation was seen in monocytes. The activation was mainly produced by components of the bacterial cell and no exo-products, because heat-killed and live bacteria caused high activation of caspase-1 as well as cytokine production, whereas the bacterial supernatant elicited minor effect. The response among different BDs varied significantly, almost fivefold. In addition, P. acnes of various origins showed considerable variation, however, the commensal isolates showed a stronger response compared with the invasive. In conclusion, although regarded as a harmless commensal of the skin, P. acnes strongly activates the inflammasome of human peripheral neutrophils.

Place, publisher, year, edition, pages
Hoboken, USA: Wiley-Blackwell, 2013
Keywords
Propionibacterium acnes, anaerobic bacteria, innate immunity, inflammasome, leukocytes, prostate.
National Category
Biomedical Laboratory Science/Technology
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-30113 (URN)10.1111/apm.12035 (DOI)000320614200009 ()23278288 (PubMedID)2-s2.0-84879497934 (Scopus ID)
Note

Funding Agencies:

Research Committee of the County Council of Orebro 

Nyckelfonden at Orebro University Hospital 

King Gustaf V Memorial Foundation 

Available from: 2013-08-02 Created: 2013-08-02 Last updated: 2018-09-11Bibliographically approved
3. Staphylococcus aureus-mediated caspase-1 activation in human neutrophils: a role for Panton-Valentine Leukocidin?
Open this publication in new window or tab >>Staphylococcus aureus-mediated caspase-1 activation in human neutrophils: a role for Panton-Valentine Leukocidin?
(English)Manuscript (preprint) (Other academic)
National Category
Biomedical Laboratory Science/Technology
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-30115 (URN)
Available from: 2013-08-02 Created: 2013-08-02 Last updated: 2017-10-17Bibliographically approved
4. Cytokine profile in a cohort of healthy blood donors carrying polymorphisms in genes encoding the nlrp3 inflammasome
Open this publication in new window or tab >>Cytokine profile in a cohort of healthy blood donors carrying polymorphisms in genes encoding the nlrp3 inflammasome
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Background: The NLRP3 inflammasome has been recognized as one of the key components of the innate immunity by sensing a diversity of insults. Inflammasome activation results in the maturation of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18. Increased production of IL-1β is found in patients with gain-of-function polymorphisms in genes encoding the NLRP3 inflammasome. Since approximately 5% of the Swedish population are heterozygote carriers of these combined gene variants, their impact on inflammasome status and a relationship on disease development is therefore highly relevant to study. The present study investigates levels of inflammasome-produced cytokines as a measure of inflammasome activation in healthy individuals carrying Q705K polymorphism in the NLRP3 gene combined with C10X in the CARD8 gene.

Materials and Methods: Genotyping of 1006 healthy blood donors was performed for the polymorphisms Q705K in the NLRP3 and C10X in the CARD8 genes. IL-1β, IL-18, IL-33, as well as a number of other pro-inflammatory cytokines, were analyzed by Luminex or ELISA in plasma from individuals carrying the polymorphisms and in age and gender matched noncarrier controls.

Results & Discussion: The prevalence of the polymorphisms was in line with previous studies. Plasma levels of IL-1β and IL-33 were elevated among carriers of combined Q705K/C10X polymorphisms compared to controls, whereas no difference was found for IL- 18 and the other cytokines measured. These data suggest that these combined polymorphisms creates inflammasomes with increased basal activation state, which might provide a more favourable innate immune response. In spite of this, it could also represent the mechanisms by which the inflammatory loop is triggered into a long-term inflammatory phenotype.

Keywords
Auto-inflammation, Cytokines, Inflammasome, Interleukin-1β, Leukocytes
National Category
Medical and Health Sciences Immunology
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-32006 (URN)
Available from: 2013-10-15 Created: 2013-10-14 Last updated: 2017-10-17Bibliographically approved
5. C10X polymorphism in the CARD8 gene is associated with bacteraemia
Open this publication in new window or tab >>C10X polymorphism in the CARD8 gene is associated with bacteraemia
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Introduction:: The NLRP3 inflammasome is an intracellular multi-protein complex that triggers caspase-1 mediated maturation of interleukin-1β (IL-1β); one of the most potent mediators of inflammation and a major cytokine produced during severe infections, like sepsis. However, the excessive cytokine levels seem to stage for tissue injury and organ failure, and high levels of IL-1β correlates with severity and mortality of sepsis. Instead, recent data suggest caspase- 1 to function as a guardian against severe infections. CARD8 has been implied to regulate the synthesis of IL-1β via interaction to caspase-1. In recent years, polymorphism of CARD8 (C10X) per se or in combination with NLRP3 (Q705K) has been implicated with increased risk of inflammation. The aim was to investigate the correlation of these polymorphisms with severe blood stream infection.

Methods:: Human DNA was extracted from blood culture bottles that were found to be positive for microbial growth (i.e. patients with bacteraemia). Polymorphisms Q705K in the NLRP3 gene and C10X in the CARD8 gene were genotyped using TaqMan genotyping assay. The results were compared to healthy controls and to samples from patients with negative cultures.

Results:: The polymorphism C10X was significantly over-represented among patients with bacteraemia as compared to healthy controls, whereas patients with negative blood culture were not associated with a higher prevalence. No association was observed with polymorphism Q705K of NLRP3 in eithergroup of patients.

Conclusions:: Patients carrying polymorphism C10X in the CARD8 gene are at increased risk of developing bacteraemia and severe inflammation.

Keywords
bacteraemia, blood culture, gene variants, infection, inflammasomes, inflammation, innate immunity, leukocytes, polymorphisms, sepsis
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-32015 (URN)
Available from: 2013-10-15 Created: 2013-10-15 Last updated: 2017-10-17Bibliographically approved

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