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Caspase 1 involvement in human monocyte lysis induced by Actinobacillus actinomycetemcomitans leukotoxin
Umeå University, Faculty of Medicine, Department of Odontology, Periodontology.
Umeå University, Faculty of Medicine, Department of Odontology, Periodontology.
Umeå University, Faculty of Medicine, Department of Odontology, Oral Microbiology.
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2003 (English)In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 71, no 8, 4448-4455 p.Article in journal (Refereed) Published
Abstract [en]

Actinobacillus actinomycetemcomitans, an oral bacterium implicated in the etiology of periodontal diseases, produces a leukotoxin that selectively lyses primate neutrophils and monocytes, the major populations of defense cells in the periodontium. Though lysis requires expression of the receptor lymphocyte function-associated molecule 1 (LFA-1) on the cell surface, not all LFA-1-expressing leukocyte populations are equally susceptible to the toxin. In this study, the susceptibility of human leukocytes to leukotoxin-induced lysis is compared to their expression of LFA-1 and the activity of caspase 1. Cytolysis was determined by the activity of lactate dehydrogenase released from peripheral human leukocytes after 1-h exposure to leukotoxin. Monocytes were lysed at leukotoxin concentrations of > or = 5 ng/ml, while the corresponding values for neutrophils and lymphocytes were approximately 10 times greater. Similar LFA-1 expression was found in all susceptible cell populations irrespective of their degree of sensitivity to the toxin. Exposure of monocytes to leukotoxin increased their caspase 1 activity about fivefold within 10 to 20 min. Presence of the caspase 1 inhibitor Ac-YVAD-CMK significantly blocked the leukotoxin-induced lysis of monocytes only. At sublytic concentrations, leukotoxin induced no apoptotic activity in monocytes, as revealed by the lack of caspase 3 activation and DNA fragmentation. Monocytes are the most lysis-sensitive leukocytes for A. actinomycetemcomitans leukotoxin. Their lysis by this toxin depends on caspase 1 activation and proceeds through a process that differs from classical apoptosis.

Place, publisher, year, edition, pages
2003. Vol. 71, no 8, 4448-4455 p.
URN: urn:nbn:se:umu:diva-6485DOI: 10.1128/IAI.71.8.4448-4455.2003PubMedID: 12874324OAI: diva2:146154
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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