Experimental Study Of Local Injection Of M1 Macrophage Extracellular Vesicles Promoting Expansion Of The Midpalatal Suture In Rats

Objective: Midpalatal suture expansion is a widely used treatment in orthodontic practice such as maxillary transverse deficiency.The effect of modulating the local inflammatory response of the suture on midpalatal suture expansion is unclear.M1 macrophages,a subtype of macrophages that mediates inflammation,secrete extracellular vesicles that promote transcriptional activation of the inflammatory pathway NF-κB and enhance the local immune inflammatory response.This study aimed to investigate the effect of the local application of M1 macrophage-derived extracellular vesicles on the expansion of rat midpalatal sutures and to explore the preliminary mechanism.Methods: The M1/M2 macrophage was identified by flow cytometry and RT-q PCR using a cytokine-stimulated RAW264.7 cell line.Extracellular vesicles were isolated and characterized by immunoblotting,nanoparticle analysis,and transmission electron microscopy.Twenty-four rats were selected at 6-7 weeks of age and randomly divided into a control group,expansion group,M1 extracellular vesicle treatment group,and M2 extracellular vesicle treatment group.Expander was made of 0.016 in.Nickel-titanium wire with a force of 70 g for 7 days.M1/M2 macrophage extracellular vesicles were injected locally into the midpalatal suture,and the expansion group was injected with an equal volume of 0.9% saline,while the control group did not apply any expansion force.Bone volume changes in the region of interest(ROI)of each group were detected by Micro-CT,and the histological morphology,osteoclast number,RANKL,P65,and BMP-2 expression of the midpalatal suture were detected by hematoxylin-eosin staining and immunohistochemical staining in each group.Results: The distance between palatal bones was significantly wider in the M1 macrophage extracellular vesicle-treated group(P < 0.001),and bone volume in the midpalatal suture region was significantly lower than in the other three groups,while the proportion of soft tissue increased(P < 0.05).Histological examination showed that compared with the other three groups,the M1 macrophage extracellular vesicle-treated group had a large amount of transverse fibrous tissue widening and remodeling in the midpalatal suture region,a significant increase in the number of osteoclasts and RANKL expression(P < 0.01),and a significant increase in the level of P65 protein expression(P< 0.001).Conclusion: Local injection of M1 macrophage extracellular vesicles may promote local inflammatory response by regulating the NF-k B p65 pathway,while promoting osteoclastogenesis and RANKL expression,increasing bone turnover activity,and improving the clinical efficacy of midpalatal suture expansion...