Metabolic Remodeling, Inflammasome Activation, and Pyroptosis in Macrophages Stimulated by Porphyromonas gingivalis and Its Outer Membrane Vesicles

Abstract

Porphyromonas gingivalis is one of the bacterial species most closely associated with periodontitis and can shed large numbers of outer membrane vesicles (OMVs), which are increasingly thought to play a significant role in bacterial virulence and pathogenicity. Macrophages are amongst the first immune cells to respond to bacteria and their products, so we sought to directly compare the response of macrophages to P. gingivalis or its purified OMVs. Macrophages stimulated with OMVs produced large amounts of TNFα, IL-12p70, IL-6, IL-10, IFNβ, and nitric oxide compared to cells infected with P. gingivalis, which produced very low levels of these mediators. Both P. gingivalis and OMVs induced a shift in macrophage metabolism from oxidative phosphorylation (OXPHOS) to glycolysis, which was supported by enhanced lactate release, decreased mitochondrial oxygen consumption with reduced spare respiratory capacity, as well as increased mitochondrial reactive oxygen species (ROS) production. Corresponding to this metabolic shift, gene expression analysis of macrophages infected with P. gingivalis or stimulated with OMVs revealed a broad transcriptional upregulation of genes critical to glycolysis and a downregulation of genes associated with the TCA cycle. Upon examination of inflammasome signaling and pyroptosis it was found that P. gingivalis did not activate the inflammasome in macrophages as the mature forms of caspase-1, IL-1β, and IL-18 were not detected and there was no extracellular release of lactate dehydrogenase (LDH) or 7-AAD staining. In comparison, macrophages stimulated with OMVs potently activated caspase-1, produced large amounts of IL-1β, IL-18, released LDH, and were positive for 7-AAD indicative of pyroptotic cell death. These data directly quantitate the distinct effects of P. gingivalis and its OMVs on macrophage inflammatory phenotype, mitochondrial function, inflammasome activation, and pyroptotic cell death that may have potential implications for their roles in chronic periodontitis.

Porphyromonas gingivalis triggers NLRP3‐mediated inflammasome activation in macrophages in a bacterial gingipains‐independent manner

“…Two reports showed that outer membrane vesicles (OMVs), spherical membranous structures at the outer cell membrane, have the ability to trigger inflammasome activation . To examine whether or not OMVs are involved in inflammasome activation, PMA‐differentiated THP‐1 cells were treated with bacterial culture supernatants of an isogenic mutant KDP361 lacking PorV (Δ porV ) , which not only shows mutation of one of the components of T9SS, but also downregulation of the amount of OMVs to less than 50%, as compared to WT bacteria .…”

“…In this study, secreted/released factors from bacteria were shown to be involved in triggering NLRP3 inflammasome. Although what triggers inflammasome awaits further investigation at the molecular level, one possible candidate may be OMVs, the spherical membrane structures . The macrophages stimulated with OMVs from WT P. gingivalis produced inflammatory cytokines and nitric oxide, and inflammasome activation was triggered .…”

Danger signals in oral cavity-related diseases

“…A number of periodontal pathogens, including Porphyromonas gingivalis , Fusobacterium nucleatum , Aggregatibacter actinomycetemcomitans, Treponema denticola , and Tannerella forsythia are capable of eliciting an Il‐1β response from immune cells as well as from non‐immune cells including gingival epithelial cells and fibroblasts . In addition to whole bacterial interactions, recent studies have found outer membrane vesicles from P. gingivalis, T. denticola , and T. forsythia are capable of inducing the inflammasome and IL‐1β release . Although the majority of studies indicate signaling through the intracellular NLRP3 inflammasome is required, the exact bacterial components required to fully activate complement and IL‐1β release, or if DAMPs such as ATP are additionally required, remains uncertain.…”

Impact of nitric oxide's bidirectional role on glaucoma: focus on Helicobacter pylori–related nitrosative stress

“…Beyond parthanatos and mitochondrial permeability transition, other mechanisms involved in neuronal death include intrinsic and extrinsic pathways of the apoptotic process, autophagy, phagoptosis (phagocytosis), oncosis, necroptosis, ferroptosis, sarmoptosis, autosis, autolysis, paraptosis, and pyroptosis . NO has been implicated in all these mechanisms . Figure charts the pathways associated with NO concentration fluctuations.…”