Enterococcus Faecalis activates NLRP3 inflammasomes leading to increased interleukin-1 beta secretion and pyroptosis of THP-1 macrophages

Objectives: Enterococcus faecalis is the bacterial species closely related to persistent infection in root canals. Interleukin-1 beta (IL-1 beta) is the most commonly detected proinflammatory cytokine in periapical granulation tissue and plays a critical role in host defenses against microbial infection. The synthesis and secretion of IL-1 beta are mediated mainly by Toll-like receptors and inflammasome activation. The previous study found that the nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) and the absent in Melanoma 2 (AIM2) inflammasomes are positively expressed in periapical granulation tissue. The aim of this study was to investigate the pathogenicity of E. faecalis and the molecular mechanisms of IL-1 beta secretion by THP-1 macrophages infected with E. faecalis. Methods: The IL-1 beta and lactate dehydrogenase (LDH) levels induced by E. faecalis were investigated with enzymelinked immunosorbent assay (ELISA) kit and cytotoxicity assay kit, caspase-1 and inflammasome expression levels were investigated using real time PCR and Western blot analysis. Then the effect of caspase-1, NLRP3, adenosine triphosphate (ATP), and extracellular K+ on IL-1 beta and LDH secretion, Gasdermin-D (GSDMD) cleavage induced by E. faecalis were analyzed. Results: E. faecalis significantly increased IL-1 beta and LDH release, caspase-1 and GSDMD cleavage, and NLRP3 inflammasome activation. It also showed that IL-1 beta and LDH release, GSDMD cleavage required caspase-1 and NLRP3 activation. Furthermore, the expression and activation of caspase-1 and NLRP3 were blocked by oxidized ATP and extracellular K+. Conclusion: E. faecalis infection activated caspase-1 and the NLRP3 inflammasome to induce IL-1 beta secretion and inflammatory cell death (pyroptosis). Furthermore, the activation and expression of NLRP3 induced by E. faecalis required P2X7R and K+ efflux. This study furthers our understanding of the inflammatory response mechanism induced by E. faecalis indicates that NLRP3 may be a potential target for treatment and prevention of persistent periodontitis caused by E. faecalis.

Automated summary

This study aimed to investigate the pathogenicity of Enterococcus faecalis and the molecular mechanisms of IL-1 beta secretion by THP-1 macrophages infected with E. faecalis. The results showed that E. faecalis infection activated caspase-1 and the NLRP3 inflammasome to induce IL-1 beta secretion and inflammatory cell death (pyroptosis). Furthermore, the activation and expression of NLRP3 induced by E. faecalis required P2X7R and K+ efflux, suggesting NLRP3 may be a potential target for treatment and prevention of persistent periodontitis caused by E. faecalis.