IADR Abstract Archives
Hypoxia Enhances Macrophage Immune Response in Persistent Apical Periodontitis
Objectives: Oxygen tension varies within the infected root canal system and is expected to be at its lowest in the apical third of the root canal. This study aims to study the effect of hypoxia on macrophages infected with putative endodontic pathogens associated with persistent apical periodontitis.
Methods: Macrophage cell line RAW 264.7 was infected with heat-killed Enterococcus faecalis, Streptococcus mitis, Streptococcus oralis, Fusobacterium nucleatum, Treponema denticola, and Tannerella forsythia, and subsequently incubated under normoxic and hypoxic conditions for 24 hours. Viability of macrophages following bacterial infection was determined through the MTS assay. Metabolic activity of macrophages under normoxic and hypoxic conditions were compared by measuring dehydrogenase activity. The immune response of macrophages to bacterial infection under normoxic and hypoxic conditions were determined by examining the expression of key pro-inflammatory cytokines, i.e. Interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), Interleukin-6 (IL-6), and keratinocyte chemoattractant (KC) by quantitative polymerase chain reaction (qPCR).
Results: Bacterial infection did not significantly alter the viability of macrophages under both hypoxic and normoxic conditions. Although macrophages exhibited a 4-fold lower metabolic activity under hypoxia, significant increases in the expression of all cytokines examined, with the exception of TNF-α, were observed in infected macrophages when compared to normoxia. Interestingly, macrophages infected with F. nucleatum under hypoxia displayed up to a 73-fold greater expression of cytokines, when compared to similar samples infected with other bacteria used in this study.
Conclusions: We showed for the first time that in hypoxic conditions, macrophages had reduced metabolic activity but demonstrated an increased immune response to bacterial insult. Oxygen tension is likely to influence host-microbe interactions in persistent apical periodontitis. Our research also highlighted the strong immune response elicited by F. nucleatum, and its potential role in disease pathogenesis.
Methods: Macrophage cell line RAW 264.7 was infected with heat-killed Enterococcus faecalis, Streptococcus mitis, Streptococcus oralis, Fusobacterium nucleatum, Treponema denticola, and Tannerella forsythia, and subsequently incubated under normoxic and hypoxic conditions for 24 hours. Viability of macrophages following bacterial infection was determined through the MTS assay. Metabolic activity of macrophages under normoxic and hypoxic conditions were compared by measuring dehydrogenase activity. The immune response of macrophages to bacterial infection under normoxic and hypoxic conditions were determined by examining the expression of key pro-inflammatory cytokines, i.e. Interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), Interleukin-6 (IL-6), and keratinocyte chemoattractant (KC) by quantitative polymerase chain reaction (qPCR).
Results: Bacterial infection did not significantly alter the viability of macrophages under both hypoxic and normoxic conditions. Although macrophages exhibited a 4-fold lower metabolic activity under hypoxia, significant increases in the expression of all cytokines examined, with the exception of TNF-α, were observed in infected macrophages when compared to normoxia. Interestingly, macrophages infected with F. nucleatum under hypoxia displayed up to a 73-fold greater expression of cytokines, when compared to similar samples infected with other bacteria used in this study.
Conclusions: We showed for the first time that in hypoxic conditions, macrophages had reduced metabolic activity but demonstrated an increased immune response to bacterial insult. Oxygen tension is likely to influence host-microbe interactions in persistent apical periodontitis. Our research also highlighted the strong immune response elicited by F. nucleatum, and its potential role in disease pathogenesis.