Inhibition of Dynamin-Related Protein 1-Dependent Mitochondrial Fission Ameliorates Apical Periodontitis by Attenuating NLRP3 Inflammasome-Mediated M1 Macrophage Polarisation.

Objective: The aim of this study was to determine if Dynamin-related protein 1‌ (Drp1) -dependent mitochondrial fission is involved in the pathological process of apical periodontitis and elucidate the underlying mechanisms.

Methods: Immunohistochemistry, immunofluorescence, and western blotting assessed CD86 expression, NLRP3/Caspase1/IL-1β activation, and mitochondrial dynamics-related proteins in human periapical macrophages. In vitro apical periodontitis (AP) models used macrophages stimulated with P. gingivalis LPS (Pg-LPS), pretreated with/without Drp1 inhibitor Mdivi-1 or NLRP3 inhibitor MCC950. CD86, NLRP3/Caspase1/IL-1β, mitochondrial dynamics proteins, TNF, and IL-6 were quantified via western blot, RT-qPCR, or immunofluorescence. ROS (DCFH-DA), ATP (commercial kit), mitochondrial membrane potential (JC-1), and morphology (TEM, MitoTracker/immunofluorescence) were analysed. Multimodal approaches explored Drp1-mediated mitochondrial fission and NLRP3 inflammasome-dependent M1 polarisation. Mdivi-1-treated AP models were established to dissect Drp1's role in pathogenesis.

Results: Our findings indicate that the Drp1-mediated excessive mitochondrial fission is present in human periapical lesions. Moreover, there was a positive correlation between the p-Drp1(Ser616) and the elevation of NLRP3, Cleaved-Caspase1, and CD86. In vitro experiments demonstrated that Mdivi-1 effectively inhibited the Pg-LPS induced abnormal mitochondrial fragmentation dependent on p-Drp1(Ser616), rescued mitochondrial dysfunction, and further suppressed the activation of NLRP3/Caspase1/IL1β and the expression of CD86. In vivo experiments showed that Mdivi-1 treatment could significantly alleviate the inflammatory bone erosion at the apex in a murine AP model by inhibiting macrophage polarisation and the NLRP3 inflammatory pathway.

Conclusions: Drp1-mediated excessive mitochondrial fission plays a crucial role in the development of AP by promoting NLRP3 signaling pathway-dependent macrophage M1 polarisation. Targeting Drp1 may serve as a potential therapeutic strategy to prevent AP development, highlighting its clinical translational value. Conclusions: Mdivi-1 can restore mitochondrial dynamics homeostasis in periapical macrophages, inhibit NLRP3/M1 polarisation, and reduce bone resorption. This suggests that Mdivi-1 might be a promising agent for management of AP in future.