Targeting periodontal inflammatory microenvironment to ameliorate periodontitis: A nitrogen ions implantation technique.

Periodontitis, an inflammatory disease affecting over 45 % of adults globally, causes irreversible alveolar bone loss through immune cells-mediated inflammation, thus necessitating novel regenerative therapies. This study proposes a cold plasma-based nitrogen species implantation (PBNI) technique to regulate the periodontal immune microenvironment and ameliorate periodontal inflammation. This study demonstrates that PBNI, optimizing for nitric oxide (NO) delivery, reprograms macrophage phenotype to resolve inflammation and reduce alveolar bone resorption. The linear correlation between exposure time and NO yield establishes PBNI as a tunable NO delivery approach. In mice with experimental periodontitis, PBNI increased alveolar bone volume, the effects of which were mirrored by the NO donor L-arginine. High-throughput RNA sequencing of bone marrow-derived macrophages (BMDMs) exposed to PBNI identified a cluster of differentially expressed genes responding to the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT)/AMP-activated protein kinase (AMPK) pathways significantly enriched. Mechanistically, PBNI-derived NO activates the canonical soluble guanylate cyclase/Protein Kinase G (sGC/PKG) cascade and a newly identified phosphoinositide-3-kinase adaptor protein 1 (PIK3AP1)/AKT1/AMPKα1 axis, synergistically driving the M2 macrophage repolarization. Crucially, NO scavenging or PI3K pathway inhibition abrogated M2 repolarization and alveolar bone regeneration. This study establishes a tunable NO delivery approach that coordinates sGC/PKG and PIK3AP1/AKT1/AMPKα1 signaling to reprogram periodontal inflammatory microenvironment. By resolving the mechanism of PBNI and enhancing its osteogenic efficacy through controlling NO release, we provide a clinically translatable strategy for immunomodulatory periodontal regeneration therapy.