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Cytokine responses of human gingival fibroblasts to Actinobacillus actinomycetemcomitans cytolethal distending toxin
Umeå University, Faculty of Medicine, Odontology, Oral Microbiology. Umeå University, Faculty of Medicine, Odontology, Oral Cell Biology.
Umeå University, Faculty of Medicine, Odontology, Periodontology.
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2005 (English)In: Cytokine, ISSN 1043-4666, Vol. 30, no 2, 56-63 p.Article in journal (Refereed) Published
Abstract [en]

Actinobacillus actinomycetemcomitans is implicated in the pathogenesis of localized aggressive periodontitis, and has the capacity to express a cytolethal distending toxin (Cdt). Gingival fibroblasts (GF) are resident cells of the periodontium, which can express several osteolytic cytokines. The aims of this study were a) to investigate the role of Cdt in A. actinomycetemcomitans-induced expression of osteolytic cytokines and their cognate receptors in GF and b) to determine if the previously demonstrated induction of receptor activator of NFkappaB ligand (RANKL) by A. actinomycetemcomitans is mediated by these pro-inflammatory cytokines or by prostaglandin E(2) (PGE(2)). A. actinomycetemcomitans clearly induced interleukin (IL)-6, IL-1beta, and to a minimal extent, tumor necrosis factor (TNF)-alpha mRNA expression. At the protein level, IL-6 but not IL-1beta or TNF-alpha expression was stimulated. The mRNA expression of the different receptor subtypes recognizing IL-6, IL-1beta and TNF-alpha was not affected. A cdt-knockout strain of A. actinomycetemcomitans had similar effects on cytokine and cytokine receptor mRNA expression, compared to its parental wild-type strain. Purified Cdt stimulated IL-6, but not IL-1beta or TNF-alpha protein biosynthesis. Antibodies neutralizing IL-6, IL-1 or TNF-alpha, and the PGE(2) synthesis inhibitor indomethacin, did not affect A. actinomycetemcomitans-induced RANKL expression. In conclusion, a) A. actinomycetemcomitans induces IL-6 production in GF by a mechanism largely independent of its Cdt and b) A. actinomycetemcomitans-induced RANKL expression in GF occurs independently of IL-1, IL-6, TNF-alpha, or PGE(2).

Place, publisher, year, edition, pages
2005. Vol. 30, no 2, 56-63 p.
URN: urn:nbn:se:umu:diva-18083DOI: 10.1016/j.cyto.2004.11.008PubMedID: 15804596OAI: diva2:157756
Available from: 2007-12-13 Created: 2007-12-13 Last updated: 2009-08-21Bibliographically approved
In thesis
1. Cellular and molecular responses of periodontal connective tissue cells to Actinobacillus actinomycetemcomitans cytolethal distending toxin
Open this publication in new window or tab >>Cellular and molecular responses of periodontal connective tissue cells to Actinobacillus actinomycetemcomitans cytolethal distending toxin
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Actinobacillus actinomycetemcomitans is present in elevated proportions and numbers in dental bacterial biofilms of patients with localized aggressive periodontitis. This variant of periodontal disease, occurring in adolescents and young adults, is characterized by rapid and severe destruction of the connective tissues and bone supporting the teeth, eventually culminating in tooth loss. The cytolethal distending toxin (Cdt) is a newly discovered bacterial protein toxin, uniquely present in A. actinomycetemcomitans among all known to-date oral bacterial species. The Cdt has the capacity to inhibit mammalian cell growth, but its putative role in the pathogenesis of the disease is unclear. The aim of this in vitro work has been to study the effects of A. actinomycetemcomitans on periodontal connective tissue cell cultures, and to evaluate the possible involvement of its Cdt.

A. actinomycetemcomitans inhibited the proliferation of gingival and periodontal ligament fibroblasts, as a result of a combined arrest at the G1 and G2/M phases of the cell cycle. This growth inhibition was non-lethal and the cells remained metabolically active, although their DNA synthesis was reduced. The intoxicated cells exhibited increased size and irregular structure, characterized by distension and elongation. This cellular enlargement occurred in both G1 and G2/M phase arrested cells. The Cdt of A. actinomycetemcomitans was responsible for the observed growth inhibition, as well as the concomitant morphological alterations.

The possible induction of inflammatory cytokines related to bone resorption was investigated in response to A. actinomycetemcomitans, and the involvement of Cdt was evaluated. Extensive focus was given to the study of receptor activator of NF-κB ligand (RANKL) expression, a membrane-bound ligand that signals osteoclast progenitors to differentiate and fuse into mature osteoclasts, activating bone resorption. It was demonstrated that A. actinomycetemcomitans induced RANKL mRNA and protein expression in the cells studied, but did not affect the expression of its decoy receptor, osteoprotegerin. This induction was solely attributed to its Cdt, as demonstrated by the use of a cdt-knockout A. actinomycetemcomitans strain, purified recombinant Cdt, and antibodies blocking the Cdt. In addition, this event was not mediated by pro-inflammatory cytokines known to stimulate RANKL. Interleukin-6 mRNA and protein expression were also enhanced by A. actinomycetemcomitans, but Cdt had limited involvement in this enhancement.

In conclusion, two distinct mechanisms by which A. actinomycetemcomitans Cdt may be involved in the pathogenesis of localized aggressive periodontitis are proposed. Firstly, the growth arrest of the resident fibroblasts may impair the physiological connective tissue remodelling equilibrium and lead to connective tissue attachment loss. Secondly, the induction of RANKL by these cells, residing in the proximity of the alveolar bone, may locally stimulate osteoclastogenesis and promote alveolar bone resorption. This work also provides further insights to the understanding of Cdt mechanisms of action, contributing to the global characterization of the toxin’s virulence.

Place, publisher, year, edition, pages
Umeå: Odontologi, 2004. 49 p.
Umeå University odontological dissertations, ISSN 0345-7532 ; 88
Medical sciences, Actinobacillus actinomycetemcomitans, cytolethal distending toxin, periodontal connective tissue cells, localized aggressive periodontitis, growth arrest, bone resorption, receptor activator of NF-κB ligand, osteoprotegerin, pro-inflammatory cytokines, MEDICIN OCH VÅRD
National Category
Medical and Health Sciences
Research subject
urn:nbn:se:umu:diva-345 (URN)91-7305-746-0 (ISBN)
Public defence
2004-12-10, Sal 933, 3A, Norrlands Universitessjukhus, Umeå, 13:00 (English)
Available from: 2004-11-01 Created: 2004-11-01 Last updated: 2009-08-24Bibliographically approved

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