Exogenous Nitric Oxide Ameliorates Macrophage-Mediated Periodontal Inflammation

The nitric oxide (NO) is a reactive nitrogen species closely related to the periodontitis treatment. This study aims to investigate the roles and regulatory mechanisms of exogenous NO from cold atmospheric plasma (CAP) in modulating periodontal inflammation and alveolar bone regeneration. A mouse experimental periodontitis model was constructed by suture ligature. The effects of helium-nitrogen CAP (HeO2-CAP) and helium-nitrogen CAP (HeN2-CAP) on inflammatory state was assessed by microcomputed tomography, hematoxylin & eosin and immunohistochemical staining. Flow cytometry detected the immune cells infiltration in mice gingival tissues. The RNA sequencing data from GEO dataset was used to analyze the differential genes associated with M1 macrophage polarization. The bone marrow-derived macrophages (BMDMs) were isolated to detect the expression of signaling pathways in M1 macrophage and the effects of CAP on macrophage repolarization. P < 0.05 was considered as statistically significant. HeN2-CAP significantly reduced inflammatory indexes, increased alveolar bone volume and decreased macrophage infiltration in gingival tissues of mice with periodontitis. The unfolded protein response (UPR) signal pathways were activated in M1-BMDMs stimulated by lipopolysaccharide (LPS) according to the data of RNA sequencing and in vitro experiments. The NO from HeN2-CAP increased the expression of M2 macrophage markers and alveolar bone regeneration of mice, as well as down-regulated the protein expression of Inositol requiring enzyme 1α (IRE1α) pathways in M1 BMDMs. The NO from HeN2-CAP reducing M1 macrophage polarization through inhibiting the IRE1α pathway, thus alleviating periodontal inflammation and enhancing alveolar bone regeneration.