Mechanistic Study Of Drp1 Modulating Microglial Proinflammatory Factor Expression Induced By Porphyromonas Gingivalis Lipopolysaccharides

Objective: Periodontitis,a common chronic inflammatory oral disease in humans,has been recognized as a key factor in the pathogenesis of neurodegenerative diseases such as Alzheimer’s disease(AD).Porphyromonas gingivalis lipopolysaccharides(P.g.LPS),a primary factor in periodontitis,can destroy the blood-brain barrier and enter the brain of patients with neurodegenerative diseases and activate microglia,releasing inflammatory factors and mediating neuroinflammation and neurotoxicity.Mitochondrial dynamics play a crucial role in driving microglial metabolism and immune response,and are involved in the progression of neurodegenerative diseases such as AD.This study investigates the mechanism by which P.g.LPS-induced changes of Dynamin-related protein 1(Drp1)in microglia,result in increased inflammation and acceleration of the neurodegenerative disease process.The results of this study provide a theoretical foundation for the relationship between periodontitis and neuroinflammation and neurodegenerative diseases such as AD.Methods:(1)The effects of P.g.LPS on microglia cells were investigated to establish a model of microglial inflammation mediated by periodontitis-associated factors.Western blot was used to detect expression of pro-inflammatory mediators TNF-α,NO,IL-1β.(2)Western blot was used to detect expression of Drp1 oligomerization,phosphorylation at Ser616,as well as other mitochondrial dynamics-related molecular expression.(3)The influence of silenced Drp1 on TNF-α and IL-1β expression in microglia cells treated with P.g.LPS was determined by Western blot.(4)Proteomics and bioinformatics methods were utilized to analyze the potential molecular mechanisms of Drp1 in P.g.LPS-treated microglia cells.(5)The influence of silenced Drp1 on Acod1 expression in P.g.LPS-treated microglia cells was investigated by Western blot and RT-q PCR.(6)The effects of silenced Acod1 on Drp1 expression and phosphorylation in P.g.LPS-treated microglia cells were analyzed using Western blot.(7)Co-IP and proteomics methods were used to explore the potential interaction mechanisms between Drp1 and Acod1.Results:(1)P.g.LPS activates the expression of pro-inflammatory mediators and neurotoxic factors in microglia;(2)P.g.LPS promotes Drp1 oligomerization in microglial cells and enhances Drp1 phosphorylation at the division site Ser616 and regulating the mitochondrial dynamics participated by Drp1 in microglia;(3)Silencing Drp1 significantly decreases the expression of pro-inflammatory factors in microglia stimulated by P.g.LPS;(4)Silencing Drp1 significantly down-regulates the up-regulation of Acod1 expression in microglia induced by P.g.LPS;(5)Silencing Acod1 has no significant effect on Drp1 expression and phosphorylation in microglia stimulated by P.g.LPS;(6)The binding of Drp1 to Acod1 increased in microglia stimulated by P.g.LPS.Conclusion: P.g.LPS can regulate the mitochondrial dynamics of Drp1 in microglia and promote the expression of inflammatory factors in microglia.In this process,Drp1 promotes the up-regulation of Acod1 and increases the binding to Acod1 protein.