Porphyromonas gingivalis outer membrane vesicles induce an activated but permissive neutrophil phenotype.

INTRODUCTION: Porphyromonas gingivalis (Pg) is a keystone periodontal pathogen linked to adverse pregnancy outcomes that releases outer membrane vesicles (OMV) capable of modulating innate immune responses; however, whether PgOMV reprogram neutrophil antimicrobial and metabolic functions, and how the bacterial oxidative environment shapes these effects, remained unknown. METHODS: To address this, we evaluated the impact of PgOMV on human peripheral neutrophil activation, NET formation, metabolic parameters, and intracellular bacterial internalization and persistence, as well as their indirect effects on trophoblast migration. We further compared responses to OMV derived from bacteria grown under basal versus oxidative stress conditions (H2O2-PgOMV). RESULTS: PgOMV induced neutrophil activation, as evidenced by increased ROS production and CD11b surface expression. Despite this activation, PgOMV did not trigger NET formation and significantly reduced extracellular DNA through DNase activity, as confirmed using PMA-induced NET preparations. PgOMV stimulation increased glucose uptake and lipid droplet accumulation, reduced lactate release, and decreased both mitochondrial mass and membrane potential. Neutrophils preconditioned with PgOMV displayed significantly greater Pg intracellular internalization and persistence relative to untreated controls, and conditioned media from PgOMV-stimulated neutrophils impaired HTR-8/SVneo trophoblast migration. Comparison of basal and H2O2-PgOMV revealed that both preparations similarly activated neutrophils, but H2O2-PgOMV did not alter glucose uptake or lactate release, and neutrophils preconditioned with H2O2-PgOMV showed significantly greater P. gingivalis internalization and persistence beyond that seen with basal PgOMV. CONCLUSIONS: Together, these findings indicate that PgOMV induce an activated but permissive neutrophil phenotype characterized by activation, impaired extracellular antimicrobial responses, altered metabolic-associated parameters, and increased susceptibility to intracellular bacterial survival and that oxidative stress further enhances bacterial persistence. The impairment of trophoblast migration by PgOMV-conditioned neutrophil media supports an indirect, neutrophil-mediated mechanism through which periodontal-derived vesicles may contribute to placental dysfunction, with implications for the link between periodontitis and adverse pregnancy outcomes.