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AIM The purpose of this study was to elucidate the suppressive effect of high-frequency pulsed diode laser irradiation on bone resorption and its biological effects for gene expression and microbiome composition on the gingival tissue in ligature-induced periodontitis in mice. MATERIALS AND METHODS Ligating ligature around the teeth and/or laser irradiation was performed on the gingival tissue in mice as follows: Co (no ligature and no laser irradiation), Li (ligation without laser irradiation), La (no ligature but with laser irradiation), and LiLa (ligation with laser irradiation). Bone resorption was evaluated using micro-computed tomography. RNA-seq analysis was performed on gingival tissues of all four groups at 3 days post ligation. The differences in microbial composition between Li and LiLa were evaluated based on the number of 16S rRNA gene sequences. RESULTS Bone resorption caused by ligation was significantly suppressed by laser irradiation. RNA-seq in Co and La gingival tissue revealed many differentially expressed genes, suggesting diode laser irradiation altered gene expression. Gene set enrichment analysis revealed mTORC1 signaling and E2F target gene sets were enriched in gingival tissues both in La and LiLa compared to that in Co and Li, respectively. The amount of extracted DNA from ligatures was reduced by laser irradiation, and bacterial network structure was altered between the Li and LiLa. CONCLUSIONS High-frequency pulsed diode laser irradiation showed biological effects and suppressed bone resorption in ligature-induced periodontitis. This article is protected by copyright. All rights reserved.