Novel Apoptosis-Inducing Activity in Bacteroides forsythus : a Comparative Study with Three Serotypes of Actinobacillus actinomycetemcomitans

Abstract

Bacteroides forsythus, which has been reported to be associated with periodontitis but has not been recognized as a key pathogen, was found to induce cytolytic activity against HL-60 and other human leukemic cells. This cytolytic activity was demonstrated according to three different criteria: (i) loss of both mitochondrial membrane potential and membrane integrity in cells treated with bacterial extracts and then with Rh123 and propidium iodide, respectively, as demonstrated by flow cytometry; (ii) damage to cytoplasmic membrane, as revealed by scanning electron microscopy (SEM); and (iii) DNA ladder formation and activation of caspase-3. These results indicate that B. forsythus produced an apoptosis-inducing factor(s) found to be composed of protein as judged by heat and trypsin sensitivity. In addition to extracts from B. forsythus, the culture supernatant of this bacterium has the ability to induce a cytolytic effect against peripheral white blood cells, especially lymphocytes. For comparison with B. forsythus, the same analyses were applied to two strains with different serotypes of Actinobacillus actinomycetemcomitans, serotypes a (ATCC 43717) and c (ATCC 43719), in addition to previously reported apoptosis-inducing serotype b (ATCC 43718), which was used as a positive control. The strains of A. actinomycetemcomitans serotypes a and b induced apoptosis in HL-60 cells as judged by the above three criteria but to a slightly lesser extent than did B. forsythus, while the serotype c strain produced apoptosis to a negligible extent. Detailed SEM images showed that the A. actinomycetemcomitans serotype a strain induced large-pore formation and the serotype b strain produced small pores with typical blebbing, while B. forsythus induced severe membrane ruffling. Further DNA ladder formation and caspase-3 activation were observed in the serotype a and b strains but not in the serotype c strain. The present paper is the first report of a protein factor(s) from B. forsythus and the A. actinomycetemcomitans serotype a strain which induces apoptotic cell death.Periodontitis, one of the most common human infectious diseases, is an acute or chronic infectious condition that can result in the inflammatory destruction of periodontal tissues such as periodontal ligaments and alveolar bone. Furthermore, recent studies have indicated that periodontitis might contribute to serious systemic diseases, such as cardiovascular diseases (18). Among more than 300 species of bacteria in the oral cavity, Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans are the major periodontopathic bacteria, and several virulence factors related to the pathogenesis of periodontitis have been reported (reviewed in references 3a, 8, and 26). In addition to these two bacteria, Bacteroides forsythus, a gramnegative, anaerobic, and fusiform bacterium (24), has recently been recognized as one of the agents associated with periodontitis. It has been reported that the presence of B. forsythus in subgingival flora was significantly associated ...

Beyond Good and Evil in the Oral Cavity: Insights into Host-Microbe Relationships Derived from Transcriptional Profiling of Gingival Cells

“…Among other effectors of the apoptotic response, the products of p53 and Bcl-2 proteins have been shown to play fundamental roles (Handfield et al 2005;Mao et al 2007;Tonetti Cortellini and Lang 1998a;Vaux Cory and Adams 1988). Emerging evidence supports the concept that bacterium-modulated apoptosis is a relevant phenomenon in the pathogenesis of periodontal diseases (Arakawa et al 2000;Bantel et al 2005a;Chen and Zychlinsky 1994;Graves Jiang and Genco 2000;Jarnbring et al 2002;Kato et al 1995;Tonetti Cortellini and Lang 1998a;Tonetti Cortellini and Lang 1998b;Vitkov Krautgartner and Hannig 2005;Wang et al 1999).…”

Biological Activities ofBacteroides forsythusLipoproteins and Their Possible Pathological Roles in Periodontal Disease

“…The virulence factors that have been identified so far are a trypsin-like protease (34,60), a sialidase (21), N-benzoyl-Val-Gly-Arg-p-nitroanilide-specific protease encoded by the prtH gene (47), and a cell surface-associated protein of B. forsythus which is involved in adhesion to fibronectin and fibrinogen (22,50). Recently, Arakawa et al have reported that a proteinous factor(s) from this bacterium is able to induce apoptosis in a human myeloid leukemia cell line, HL-60 (1).…”

Proteomic and Bioinformatic Profile of Primary Human Oral Epithelial Cells

“…Infected gingival epithelial cells attempt to eliminate challenging micro-organisms by inducing cell death to protect neighboring cells from infection 29 . Emerging evidence supports the concept that bacterially-modulated apoptosis is a relevant process in the pathogenesis of periodontal diseases 28, 30–32 . Since the oral epithelium is constantly challenged by mixed bacterial flora, it is hypothesized that a high rate of epithelial cell turnover enables greater bacterial clearance and thus limits the invasion of the gingival tissues 28, 31, 33 .…”