TNFα expression by Porphyromonas gingivalis‐stimulated macrophages relies on Sirt1 cleavage

Abstract

Objective Periodontitis is one the most common chronic inflammatory conditions, resulting in destruction of tooth‐supporting tissues and leading to tooth loss. Porphyromonas gingivalis activates host macrophages to secrete pro‐inflammatory cytokines and elicit tissue damage, in part by inducing NF‐kappa‐B transactivation. Since NFκB transactivation is negatively regulated by the Nicotinamide adenine dinucleotide (NAD)‐dependent deacetylase enzyme Sirt1, we sought to assess if RAW264.7 macrophages exposed to P. gingivalis demonstrate impaired Sirt1 activity, to ultimately induce a pro‐inflammatory response. Methods RAW264.7 macrophages were incubated with heat‐ killed P. gingivalis for 2, 4, 8, and 24 h. Stimulated RAW264.7 were assessed for TNFα expression via PCR, ELISA, and ChIP analysis. Following the activation of RAW264.7 macrophages, immunoblot analysis was executed to detect modifications in Sirt1 and the NFκB subunit RelA that is essential for NFκB transcriptional activity. Results TNFα expression was elevated 4 h after exposure to P. gingivalis. ChIP confirmed that RelA was enriched in the mouse TNFα promoter 4 h following stimulation, which correlated with the increased TNFα mRNA levels. Preceding TNFα expression, we detected Phosphoserine 536 and acetylated lysine 310 of RelA after 2 hours exposure with P. gingivalis. Moreover, reduced Sirt1 activity was associated with its cleavage in RAW264.7 protein extracts, after 2 hours of P. gingivalis exposure. Blocking TLR2/4 signaling prevented Sirt1 cleavage, loss of deacetylase activity, and TNFα secretion, while co‐administering CA074Me (a cathepsin B inhibitor) with P. gingivalis reduced RelA promoter enrichment, resulting in impaired TNFα expression. Conclusions Together, the results suggest that P. gingivalis induces TNFα expression, at least in part, by enhancing cleavage of Sirt1 via a TLR‐dependent signaling circuit.

Macrophages: A communication network linking Porphyromonas gingivalis infection and associated systemic diseases

“…With their combination, they will regulate the trimethylation of histone H3, lysine 27 (H3K27me3) on promoter regions of genes that define the M2 phenotype, thus promoting M2 macrophage activation ( 25 )( Figure 1 ). In addition, P. gingivalis activates TLR2 and TLR4 and elicits the expression of TNF-α in macrophages ( 26 ). TNF-α is an anti-M2 macrophage factor that blocks M2 macrophage polarization on two levels: through its direct effects on macrophages and the indirect effects of TNF-α on IL-13 production by other innate cell types ( 17 ).…”

Macrophages in periapical lesions: Potential roles and future directions

“…Sun et al ( 40 ) found that the dominant bacteria isolated from extraradicular and intraradicular samples from persistent APs were quite different, which suggests that different bacteria play different roles in the pathogenesis of persistent PLs. Considering the fundamental roles of bacteria in the progression of PLs, in this section we summarize the functions of oral bacteria, such as Enterococcus faecalis and P. gingivalis , in the polarization of macrophages ( 41 , 42 ).…”

NLRP3 Disturbs Treg/Th17 Cell Balance to Aggravate Apical Periodontitis

“…Heat-killed P. gingivalis (HKPG) has retained the main virulence factors, like lipopolysaccharide, fimbriae, and peptidoglycan (Akira et al 2006; El-Awady et al 2015). Hence, a frequent stimulus for imitating apical periodontitis in vitro is HKPG, which maintains immunogenicity while preventing culture contamination (Meka et al 2021). Due to an inflammatory response triggered by these pathogenic components, the innate and adaptive immune systems engage in an inflammatory destruction of periapical tissues (Márton and Kiss 2000; Fujii et al 2009).…”