Treponema denticola, Porphyromonas gingivalis, and Tannerella forsythia induce cell death and release of endogenous danger signals

Abstract Objective The aim of this study was to analyze whether periodontopathogens induced inflammatory cell death and the release of diverse endogenous danger molecules in THP-1-derived macrophages. Methods The macrophages were treated with Treponema denticola , Porphyromonas gingivalis , and Tannerella forsythia . Activation of caspase-1 and caspase-4 was detected by Western blotting. Cell death of bacteria-stimulated macrophages was examined using a lactate dehydrogenase (LDH) assay and propidium iodide (PI)/annexin V (AV) staining. Levels of endogenous danger signals, including adenosine triphosphate (ATP), uric acid, heat shock protein 60 (HSP60), high-mobility group box protein 1 (HMGB1), and fibronectin in the culture supernatants were determined using an ATP bioluminescence assay kit, a uric acid assay kit, and Western blotting, respectively. Results T. denticola , P. gingivalis , and T. forsythia induced activation of caspase-1 and caspase-4. The LDH assay and PI/AV staining showed that all three pathogens induced pyroptotic cell death. All three bacteria induced release of ATP, which is an important ligand for inflammasome activation; the increase in ATP ultimately leads to caspase-1 activation. T. denticola induced release of HSP60 and fibronectin, while T. forsythia induced release of HMGB1 in addition to HSP60 and fibronectin. None of the endogenous molecules except for fibronectin were detected in P. gingivalis -infected cells, possibly due to degradation of these factors by the proteolytic activity of the bacteria. Interestingly, P. gingivalis induced uric acid release. Conclusion Inflammatory cell death and endogenous danger molecules released from cells infected with periodontopathogens may play critical roles in the pathogenesis and progression of periodontitis by augmenting immune and inflammatory responses.