| Chapter 1: IntroductionWhite spot lesions(WSLs)and enamel cracks are the two most prominent diseases that occur after orthodontic treatment and are caused by enamel demineralization from accumulated bacterial biofilms and/ or enamel damage caused by the removal of residual adhesive after bracket debonding.Inspired by the self-assembled amelogenin nanoribbons in enamel,we developed an enamel coating with a self-assembling antimicrobial peptide(AMP),D-GL13 K,to simultaneously reduce demineralization and residual adhesive.The self-assembled amphiphilic nanoribbons significantly increased the hydrophobicity of the etched enamel,which reduced the permeability of the coated enamel surfaces as desired.The antimicrobial activity of this coating was evaluated against Streptococcus mutans by colony-forming unit(CFU)counting and live/dead assays.The anti-demineralization effect was demonstrated by the reduced demineralization depth analyzed by optical coherence tomography(OCT)and the increased Vickers hardness.The coatings did not reduce the shear bond strength(SBS)but significantly reduced the adhesive remnant index(ARI)score.This bioinspired enamel coating may provide a new strategy for preventing WSLs and enamel cracks after orthodontic treatment.Chapter 2: Construction of antimicrobial peptide D-GL13 K peptide coating on enamel surfaceObjectives The antibacterial coating on the surface of tooth enamel is a method to directly change the properties of tooth enamel and reduce the demineralization of tooth enamel.In this study,the antimicrobial peptide D-GL13 K coating on the surface of tooth enamel was constructed and its physical and chemical properties were characterized.Methods: The three-dimensional structure of antibacterial peptide D-GL1 3 K recombinant molecule was self-assembled into nanofibers by alkaline solution at p H9.9.The antimicrobial peptide coating on the enamel surface was completed by using different concentrations of antimicrobial peptide D-GL13 K and different pretreatment methods.After 24 hours at room temperature,the physical and chemical properties of the antimicrobial peptide D-GL13 K coating were characterized by water contact angle(WCA)and penetration test.At the same time,the stability of the coating was analyzed by saliva hydrolysis aging test and ultrasonic vibration test.Results:(1)The results of WCA showed that the natural enamel surface became very hydrophilic after acid etching.With the increase of D-GL13 K concentration or the addition of coatings,the hydrophobicity of the enamel surface was significantly increased,but the dehydration of the enamel surface pretreated by different methods had no significant effect on its hydrophobicity.(2)The results of penetration test showed that D-GL13 K coating had anti-methylene blue penetration effect.(3)The results of WCA test after salivary hydrolysis test showed that WCA decreased,but the enamel after coating still showed hydrophobicity,indicating that D-GL13 K had anti-proteolytic effect.(4)After the ultrasonic vibration test,the WCA decreased and the permeability increased,but the coated enamel surface still showed hydrophobicity and anti-permeability,indicating that D-GL13 K had anti-mechanical degradation effect.Conclusion: The antimicrobial peptide D-GL13 K on the enamel surface constructed in this study could form a hydrophobic coating on the surface of acid etched enamel.The coating method was simple,had anti-penetration effect,and was stable under protein degradation and mechanical stimulation.Chapter 3: Biocompatibility of antimicrobial peptide D-GL13 K enamel coatingObjective: To evaluate the cytocompatibility of antimicrobial peptide D-GL13 K coating.Methods: Rat fibroblasts(L92 9)were co cultured with 5 groups of different coated enamel slices for 2 days.The cell proliferation was quantitatively determined by CCK-8 experiment,and the cell adhesion and spreading were observed by fluorescence microscope.Results:(1)The results of CCK-8 showed that there was no significant difference in the proliferation of L929 on the enamel surface of different groups.(2)Fluorescence micrographs showed that L9 2 9 was in a healthy state under different concentrations and different coating methods.Conclusion: Antibacterial peptide D-GL13 K coating did not affect the adhesion,spreading and cell proliferation of L929,and had good cell compatibility.Chapter 4: in vitro antibacterial activity of D-GL13 K peptide coating on enamel surfaceObjective: To evaluate the antibacterial activity of antimicrobial peptide D-GL1 3 K coating in vitro and analyze the effects of different concentrations and coating methods on the antibacterial activity of antimicrobial peptide D-GL1 3 K in vitro.Methods: Streptococcus mutans was cultured for 6 hours in static anaerobic culture in vitro.Colony forming unit analysis(CFU)and live / dead bacteria staining were used to evaluate the effect of different concentrations and different coating methods on the antibacterial activity of D-GL13 K in vitro.Results:(1)The antimicrobial peptide D-GL13 K coating on the enamel surface had antibacterial activity against Streptococcus mutans.(2)The antimicrobial activity of D-GL1 3 K on enamel surface was not significantly affected by different concentrations and different coating methods.Conclusion: The antimicrobial peptide D-GL13 K coating on the enamel surface has an effective antibacterial effect on the oral cariogenic bacteria of Streptococcus mutans.Chapter 5: in vitro demineralization resistance of antimicrobial peptide D-GL13 K peptide coating on enamel surfaceObjective: Previous studies have shown that antimicrobial peptide D-GL1 3 K coating could be stable on the enamel surface and had antibacterial property.This study would further evaluate the anti-demineralization property of antimicrobial peptide D-GL13 K coating in vitro,and analyze the effects of different concentrations and coating methods on the anti-demineralization property of antimicrobial peptide D-GL13 K in vitro.Methods: After D-GL13 K coating on the enamel surface,the sterile saliva containing protease was extracted to form a biofilm on the enamel surface,and then anaerobically cultured with Streptococcus mutans for 4 days.Optical coherence tomography(OCT)and Vickers hardness analysis were used to preliminarily evaluate the effects of different concentrations and coating methods on the anti demineralization of D-GL13 K in vitro.Results:(1)The OCT curve of the antimicrobial peptide D-GL13 K coating group on the enamel surface showed that the demineralization depth was significantly reduced,and the Vickers hardness value was increased,indicating that the coating had anti-demineralization property in vitro.(2)Different concentrations and different coating methods had no significant effect on the anti-demineralization performance.Conclusion: The antimicrobial peptide D-GL13 K coating on the enamel surface had an effective anti enamel demineralization effect in the oral cariogenic environment.Chapter 6: Bond strength of brackets after coating with antimicrobial peptide D-GL13 K on enamel surfaceObjective: Previous studies had shown that antimicrobial peptide D-GL1 3 K coating had the anti-demineralization property of enamel surface,but in clinical application,the antimicrobial coating could not affect the bond strength of orthodontic brackets.Therefore,this study would further evaluate the effect of antimicrobial peptide D-GL13 K coating on the bond strength of orthodontic brackets.Methods: The roots of premolars were embedded with epoxy resin to make the crowns perpendicular to the ground plane,and then the buccal surface of the crowns was coated with antimicrobial peptide D-GL1 3 K.The orthodontic metal brackets were bonded to the coated enamel with hydrophobic adhesive.The effects of different concentrations and coating methods on the bonding strength of orthodontic treatment brackets were analyzed by Instron mechanical tester.The effect of bracket peeling on enamel surface was analyzed by ARI and SEM.Results:(1)The antimicrobial peptide D-GL13 K coating on the enamel surface did not affect the bonding strength of orthodontic brackets for clinical application.(2)The presence of D-GL13 K coating could reduce the adhesive remaining on the enamel surface.(3)SEM images showed that the coating could reduce the enamel surface damage.Conclusion: The antimicrobial peptide D-GL13 K coating on the enamel surface did not affect the bond strength of brackets in orthodontic treatment,but it could significantly reduce the enamel surface damage.Chapter 7: Conclusions and Prospect(1)In this study,the antimicrobial peptide D-GL13 K was successfully coated on the enamel surface.In vitro and in the simulated oral cariogenic environment,the D-GL1 3 K coating showed antibacterial activity and anti-demineralization property against Streptococcus mutans,and had good cell compatibility and simple clinical operation.It provided a new strategy for preventing enamel demineralization and damage in orthodontic treatment.(2)In this study,antimicrobial peptide D-GL13 K coating did not affect the bond strength of metal brackets in orthodontic treatment;After peeling off the metal brackets,the amount of residual adhesive on the enamel surface was significantly reduced,which could reduce the chair side operation time of polishing after orthodontic treatment,and at the same time protected the enamel and reduced the enamel damage caused by removing the residual adhesive on the enamel surface.(3)In this study,we need to further complete the cariogenic model animal experiment to simulate the anti-demineralization performance of antimicrobial peptide D-GL13 K coating samples in vivo under the oral environment,and carry out in-depth research from the impact of different bracket bonding strength and the analysis of enamel surface damage.At the same time,with the supply of calcium and phosphate,the remineralization potential of this biomimetic D-GL1 3 K coating will be further explored in the future work... |
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