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IL-1beta secretion induced by Aggregatibacter (Actinobacillus) actinomycetemcomitans is mainly caused by the leukotoxin
Umeå University, Faculty of Medicine, Department of Odontology, Periodontology. Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. (Aa-gruppen)
Umeå University, Faculty of Medicine, Department of Odontology. (Aa-gruppen)
Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
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2008 (English)In: International Journal of Medical Microbiology, ISSN 1438-4221, E-ISSN 1618-0607, Vol. 298, no 5/6, 529-541 p.Article in journal (Refereed) Published
Abstract [en]

Aggregatibacter (Actinobacillus) actinomycetemcomitans forms a leukotoxin that selectively lyses primate neutrophils, monocytes and triggers apoptosis in promyeloic cells and degranulation of human neutrophils. Recently, we showed that the leukotoxin causes activation of caspase-1 and abundant secretion of bio-active IL-1beta from human macrophages. In this study, we show that high levels of IL-beta correlated with a high proportion of A. actinomycetemcomitans in clinical samples from a patient with aggressive periodontitis. To determine the relative contribution of leukotoxin to the overall bacteria-induced IL-1beta secretion, macrophages were isolated from peripheral blood and exposed to different concentrations of live A. actinomycetemcomitans strains with either no, low or high production of leukotoxin. Cell lysis and levels of IL-1beta, IL-6, TNF-alpha and caspase-1 were measured by ELISA and flow cytometry. Leukotoxin was the predominant cause of IL-1beta secretion from macrophages, even in the A. actinomycetemcomitans strain with low leukotoxin production. Macrophages exposed to non-leukotoxic bacteria accumulated cytosolic pro-IL-1beta, which was secreted by a secondary exposure to leukotoxic bacteria. In conclusion, the present study shows for the first time that A. actinomycetemcomitans-induced IL-1beta secretion from human macrophages in vitro is mainly caused by leukotoxin.

Place, publisher, year, edition, pages
2008. Vol. 298, no 5/6, 529-541 p.
URN: urn:nbn:se:umu:diva-6475DOI: 10.1016/j.ijmm.2007.06.005PubMedID: 17888725OAI: diva2:146144
Available from: 2007-12-13 Created: 2007-12-13 Last updated: 2014-10-07Bibliographically approved
In thesis
1. Inflammatory cell death of human macrophages induced by Aggregatibacter actinomycetemcomitans leukotoxin
Open this publication in new window or tab >>Inflammatory cell death of human macrophages induced by Aggregatibacter actinomycetemcomitans leukotoxin
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Aggregatibacter (Actinobacillus) actinomycetemcomitans is a bacterium mainly associated with aggressive forms of periodontitis. Among its virulence factors, a leukotoxin is suggested to play an important role in the pathogenicity. Periodontal infections with strains producing high levels of the leukotoxin are strongly associated with severe disease. Leukotoxin selectively kills human leukocytes and can disrupt the local defense mechanisms. Previous studies examining the role of the leukotoxin in host-parasite interactions have mainly focused on polymorphonuclear leukocytes (PMNs). In the inflamed periodontium, macrophages play a significant role in the regulation of the inflammatory reactions and the tissue breakdown and remodeling.

Thus, the aim of this dissertation was to investigate death mechanisms of human macrophages exposed to leukotoxin.

Human lymphocytes, PMNs, and monocytes/macrophages isolated from venous blood were exposed to purified leukotoxin or live A. actinomycetemcomitans strains producing variable levels or no leukotoxin. Different target cells were characterized by their expression of cell surface molecules. Cell death and viability were studied by examining cell membrane integrity and morphological alterations. Further, processes and cellular markers involved in apoptosis and necrosis were investigated. The expression and activation of pro-inflammatory cytokines of the leukotoxin-challenged leukocytes were examined at the mRNA and protein level. The biological activity of the secreted cytokines was investigated by testing the culture supernatants in a bone resorption assay. Additionally, different intracellular signaling pathways involved in the pro-inflammatory response from the macrophages were examined.

Monocytes/macrophages were the most sensitive leukocytes for A. actinomycetemcomitans leukotoxin-induced lysis. This process in monocytes/ macrophages involved caspase-1 activation, and in addition, leukotoxin triggered abundant activation and secretion of IL-1β from these cells. The secreted IL-1β was mainly the 17 kDa bioactive protein and stimulated bone resorption. This activity could be blocked by an IL-1 receptor antagonist. When live bacteria were used, the A. actinomycetemcomitans-induced IL-1β secretion from human macrophages was mainly caused by the leukotoxin. Closer examination of the macrophages exposed to leukotoxin revealed that the induced cell death proceeded through a process that differed from classical apoptosis and necrosis. Interestingly, this process resembled a newly discovered death mechanism termed pyroptosis. The extensive leukotoxin induced IL-1β secretion did not correlate to increased levels of mRNA for IL-1β. It was mainly mediated by caspase-1 activation, since blocking it by a specific inhibitor also abolished the secretion of IL-1β. A similar pattern, but at much lower level, was seen for IL-18.

In conclusion, these results show that A. actinomycetemcomitans leukotoxin induces a death process in human macrophages leading to a specific and excessive pro-inflammatory response. Our results indicate that this novel virulence mechanism of leukotoxin may play an important role in the pathogenic potential of A. actinomycetemcomitans.

Place, publisher, year, edition, pages
Umeå: Institutionen för odontologi , 2009. 80 p.
Umeå University odontological dissertations, ISSN 0345-7532 ; 107
Aggregatibacter actinomycetemcomitans, leukotoxin, IL-1β, caspase-1, inflammatory cell death, pyroptosis
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urn:nbn:se:umu:diva-19204 (URN)978-91-7264-649-0 (ISBN)
Public defence
2009-03-27, Sal B, byggnad 1D, 9 tr, Norrlands universitetssjukhus, Norrlands universitetssjukhus, Umeå, 08:15 (English)
Available from: 2009-03-09 Created: 2009-03-05 Last updated: 2009-03-11Bibliographically approved

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Kelk, PeymanClaesson, RolfSjöstedt, AndersJohansson, Anders
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PeriodontologyClinical BacteriologyDepartment of Odontology
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