Melatonin Alleviates Periodontitis By Inhibiting Nets-Dependent Neutrophil Recruitment

This study is to investigate the roles of neutrophil extracellular traps (NETs) in periodontitis and the regulatory mechanism of melatonin on periodontitis. Gingival tissues were collected from patients with periodontitis for the expressions of NETs by immunofluorescence staining and qPCR. Transcriptome sequencing explored NETs' roles in periodontitis. In vitro, mouse bone marrow-derived neutrophils were isolated. qPCR was used to assess NETs' association with inflammatory factors (Il-1β, Il-6, Tnf-α), neutrophil chemokines (Ccl2, Ccl4, Cxcl1), and adhesion molecule (Icam-1). In vivo, Cl-Amidine was used to inhibit NETs in the ligature-induced periodontitis (LIP) mouse model (male C57BL/6J mice, 6–8 weeks age). The sleep-periodontitis association was analyzed from Public databases. LIP mice were used to explain melatonin's mechanism in periodontitis via Micro CT, histology staining, qPCR and flow cytometry. In vitro, further experiments were performed including qPCR, Immunocytochemistry staining, ELISA and ROS assay, to explain melatonin’s roles in periodontitis. Western blotting was employed to deeply explore the mechanism that melatonin inhibited NETs formation. The study showed that NETs were highly expressed in periodontitis tissues, and inhibiting NETs formation was associated with decreased neutrophils infiltration and inflammatory mediators in vitro and vivo. The close sleep-periodontitis association was revealed from Public databases, and treatment with melatonin alleviated periodontitis by inhibiting NETs-dependent neutrophil recruitment in vitro and vivo. In vitro, melatonin inhibited NETs formation via the NF-κB pathway. Melatonin mediates neutrophil recruitment via inhibition NETs, which can play protective roles on periodontal tissues.