Experimental Study Of A Cold Atmospheric Plasma Device On Canal Disinfection To Assist In Root Canal Therapy

Objective:Periapical periodontitis is an inflammatory disease in the periapical area,mainly caused by bacteria.The ability of resistant bacteria,represented by Enterococcus faecalis(E.faecalis),to form bacterial biofilms under suitable conditions and the complexity of the root canal system(lateral canals,furcations,etc.)and many other factors lead to the inability of the current stage of treatment(mechanical cleaning,drug irrigation,etc.)to completely remove bacterial biofilms.The preliminary work of our group has confirmed that cold atmospheric plasma(CAP)has a good killing effect on a variety of bacteria.Based on the previous work,the group has designed and developed a new plasma j et device,which intends to diagnose and analyze the component of active particles produced by this device,and explore the immediate and sustained killing effect of cold CAP on E.faecalis,the key causative agent of endodontic periapical disease,and to evaluate the adjunctive therapeutic effect in periapical periodontitis of Beagle dogs.Methods:A new type of plasma jet device was designed and developed to determine the active particle species by optical emission spectroscopy and the relative intensity of active particles by laser-induced fluorescence.E.faecalis biofilms were cultured on hydroxyapatite discs coated with collagen fibers for 7 days and 21 days.The samples were divided into normal saline group,cold atmospheric plasma jet group and 2%chlorhexidine group.After treatment,samples were cultured for three and seven days to obtain post-kill and re-culture samples.The samples were dehydrated and fixed for morphological observation by scanning electron microscopy.The samples were also treated with fluorescent dyes for laser confocal fluorescence intensity acquisition and three-dimensional reconstruction,followed by analysis of the dead/(live+dead)bacteria ratio.E.faecalis biofilms were cultured in human dentinal tubules for 7 days and 21 days.Grouped as described previously,fluorescent dyes were processed for laser confocal fluorescence intensity acquisition and three-dimensional reconstruction,followed by analysis of the ratio of dead/(live+dead)bacteria.Four adult Beagle dogs were subjected to periapical periodontitis modelling experiments,and after Xrays and CBCT showed apical shadows,conventional root canal therapy was performed.The dogs were randomly divided into normal saline group,cold atmospheric plasma j et group,2%chlorhexidine treatment group and negative control group.Conventional root canal preparation,conventional root canal irrigation and adjunctive treatment of the mentioned groups were performed,and finally conventional root canal filling was performed.X-rays were used for two-dimensional imaging area analysis at 30/60/90/120 days after treatment.Three-dimensional imaging volume analysis was performed using CBCT on day 120 after treatment.After the experimental animals were sacrificed,H&E staining,Masson staining and immunohistochemical staining were performed on the periapical tissues to evaluate the intervention effect of the different treatment groups on the periapical tissues of the dogs.Results:The newly developed CAP jet device operated stably and the jet temperature was close to room temperature.The spectral intensity was dominated by N2,He and O.The fluorescence intensity of O and OH was relatively high at 2～5 mm from the nozzle.For the in vitro biofilms of E.faecalis on hydroxyapatite discs,both the plasma and chlorhexidine groups significantly killed the bacteria in the biofilm compared to the saline group for both seven-day and three-week samples(P<0.05);after three-day and seven-day recultures,there was still a significant difference in the sterilization level of the two treatments compared with the normal saline group.For faecalis biofilms in human dentinal tubules,the plasma group had the ability to kill more bacteria than the chlorhexidine group(P<0.05),both for sevenday and three-week samples.X-rays and CBCT showed that the model of periapical periodontitis in beagle dogs was successfully established.At the time point of 90 and 120 days after treatment,the reduction rate of periapical periodontitis area in the plasma group was significantly different from that in the chlorhexidine group(P<0.05).On day 120 after treatment,CBCT images showed that the plasma group had the smallest reconstruction volume of apical lesion area compared with other treatment groups(P<0.05).H&E staining and Masson staining showed that the apical lesion areas in the plasma group showed bone tissue repair performance,and ALP indexes showed relatively high expression compared with other the treatment groups.Conclusion:Using CAP jet as an adjunctive therapy for root canal therapy,better healing performance was observed in imaging and histology compared with traditional root canal irrigation drugs.It is suggested that CAP jet as a root canal disinfection technology has potential application prospect to assist in root canal therapy.