| Backgrounds With the improvement of living conditions,people’s pursuit of beauty is also increasingly high.Accordingly,as a kind of dental and maxillofacial deformity,malocclusion has become a growing concern.Malocclusion not only affects the appearance,but also does harm to the growth and development of the maxilloface,and is also associated with endodontic and periodontal problems.Therefore,it is very important to treat the malocclusion in time for the psychological and physiological health of patients.However,both the traditional fixed appliances and the emerging clear aligners can keep food residues and cause poor oral hygiene due to brackets and attachments which bond to the tooth surface during treatment period.Finally,dental plaque adheres to the surface of teeth,produces acid,and results in chalky demineralization on the enamel surface.These chalky demineralization is also named white spot lesions（WSLs）.Based on nonclassical crystallization theory,amorphous calcium phosphate（ACP）has the ability to crystallize into hydroxyapatite with an orderly crystal structure.However,ACP is not stable in solution due to its poor thermodynamic stability,which means stabilizer is necessary to keep ACP stable.Previously research used polyacrylic acid（PAA）to stabilize ACP,loaded the PAA stabilized ACP（PAA-ACP）into aminefunctionalized pore-enlarging mesoporous silica nanoparticle（a MSN）to synthesize PAA-ACP@a MSN delivery system,and confirmed its remineralization ability to demineralized dentin collagen fibers.Therefore,this study aims to achieve in vitro remineralization of demineralized enamel via a mesoporous delivery system of amorphous calcium phosphate precursors（PAA-ACP@a MSN）.Part 1 Synthesis of PAA-ACP@a MSN and establishment of enamel demineralization modelExperiment 1: Preparation and characterization of PAA-ACP@a MSNPurpose:Synthesizing a MSN and loading with PAA-ACP to form PAA-ACP@a MSNMaterials and methods: In this experiment,mesoporous silicon was synthesized by de-templating method.CTAB was used as template and TMB was used as poreexpanded agent.Pore-expanded mesoporous silicon was synthesized by hydrolysis of TEOS and modified by APTES to obtain amine-functionalized pore-expanded mesoporous silica（a MSN）.Amorphous calcium phosphate（ACP）was stabilized by polyacrylic acid（PAA）to obtain PAA-ACP and PAA-ACP was loaded into a MSN to form PAA-ACP@a MSN.The PAA-ACP@a MSN paste was obtained by centrifugation.a MSN and PAA-ACP@a MSN were characterized by transmission electron microscopy（TEM）and elemental mapping analysis.Result: TEM results showed that the diameter of a MSN was about 100 nm,the hexagonal mesopores were evenly distributed on the surface of a MSN,while the mesoporous structure on PAA-ACP@a MSN was ambiguous.The elemental mapping analysis of PAA-ACP@a MSN showed that there was a uniform distribution of calcium and phosphorus elements in PAA-ACP@a MSN.Experiment 2: Establishment and evaluation of in vitro enamel demineralization modelPurpose:Establishing an in vitro enamel demineralization model and evaluating the demineralization degree.Materials and methods: Premolars extracted for orthodontic reason were collected to make enamel samples.In vitro model of enamel demineralization was established by soaking the enamel samples in demineralization solution.Before and after demineralization,the microhardness,Raman intensity and color of each enamel sample were measured by microhardness tester,micro-Raman spectrometer and spectrophotometer,and the degree of demineralization of each enamel sample was evaluated by comparing the changes of these three indexes.The changes of enamel morphology after demineralization were observed by scanning electron microscope（SEM）.Result: After demineralization,the microhardness,Raman intensity and color value of each enamel sample decreased,and the surface of the enamel became rough and uneven through SEM observation.Part 2 Evaluation of the remineralization effect of PAA-ACP@a MSNExperiment 3: Quantitative assessment of the remineralization effect of PAAACP@a MSNPurpose:Quantitatively evaluating the remineralization effect of PAA-ACP@a MSN.Materials and methods: Thirty-six demineralized enamel samples were divided into three groups: artificial saliva group（NC group,n=12）;Casein phosphopeptide calcium phosphate paste group（CPP-ACP group,n=12）;PAA-ACP@a MSN group（experimental group,n=12）.CPP-ACP paste,as a commercial remineralization agent,was used as the positive control group in this study,while the artificial saliva group was used as the negative control group.The enamel samples were remineralized by brushing different agents on surface.After remineralization process,the microhardness,Raman intensity and color values of each enamel sample were measured,and the microhardness recovery ratio,Raman intensity change ratio and color recovery ratio of each enamel sample were calculated.Result: After remineralization,the microhardness recovery ratio,Raman intensity change ratio and color recovery ratio of PAA-ACP@a MSN group were not statistically different from those of CPP-ACP group,and both two groups showed significant improvement of microhardness recovery ratio,Raman intensity change ratio and color recovery ratio when compared with the NC group.Experiment 4: Qualitative assessment of the remineralization effect of PAAACP@a MSNPurpose:Qualitatively evaluating the remineralization effect of PAA-ACP@a MSN.Materials and methods: After the quantitative evaluation,the change of the enamel morphology was observed by SEM,the crystal structure of the remineralization layer on the enamel surface was analyzed by X-Ray diffraction（XRD）,and the Ca/P atomic ratio of the remineralization layer was analyzed by Energy Dispersive Spectrometer（EDS）.Result: SEM observation showed that mineral deposits appeared on the surface of demineralized enamel after PAA-ACP@a MSN treatment.XRD analysis showed that the remineralization layer was hydroxyapatite,and EDS analysis showed that the remineralization layer had a similar calcium-phosphorus ratio to that of normal enamel. |
