Abstract:

Objective To investigate the regulatory ability and mechanism of metformin on the activation of NOD-like receptor protein 3 (NLRP3) in human periodontal ligament cells (PDLCs) by constructing a model of diabetes and periodontitis in vitro, and to provide new insights for metformin in the treatment of patients with diabetes and periodontitis. Methods Methyl thiazolyl tetrazolium assay was used to detect the cytotoxicity of metformin on human PDLCs. Human PDLCs were divided into five groups. Group A was a normal glucose concentration group (8 mmol/L glucose). Group B was a hypertonic environment group (8 mmol/L glucose and 17 mmol/L mannitol). Group C was a high concentration glucose group (25 mmol/L glucose). Group D was a high glucose environment group treated with metformin (25 mmol/L glucose+40 mmol/L metformin). Group E was a high glucose environment group treated with metformin and the AMPK inhibitor (25 mmol/L glucose+40 mmol/L metformin+ 10 μmmol/L compound C). All A-E groups were exposed to Porphyromonas gingivalis LPS at the same time. The level of IL-1β and IL-18 were detected by ELISA. The changes of proteins NLRP3, Caspase-1, ASC, AMPK, and p-AMPK were detected by western blot. Changes of mitochondrial complex Ⅰ activity were also detected. Results Metformin with a concentration not exceeding 40 mmol/L had no significant cytotoxicity on human PDLCs. Metformin treatment could significantly inhibit the inflammatory response in high glucose environments, characterized by the decrease of IL-1β, IL-18, NLRP3 and Caspase-1, and also reducing the activity of mitochondrial complex Ⅰ, while increasing the expression of p-AMPK. However, the introduction of AMPK inhibitor reversed some of the changes. Conclusion In human PDLCs, metformin reduces the activation of NLRP3 inflammasome in high glucose environment by reducing the activity of mitochondrial complex Ⅰ and activating AMPKs pathway, which provides new evidence for metformin to prevent and treat diabetes patients with periodontitis.

Key words: periodontal ligament cells, NOD-like receptor protein 3 inflammasome, metformin, mitochondrial complex Ⅰ