Dental Hard Tissue Remineralization By Using Nanogels-encapsulated Amorphous Calcium Phosphate

ObjectivesRemineralization of dental hard tissue is mainly to regenerate the dental-like hard tissue on the surface of demineralized enamel and the exposed dentinal tubules,to achieve the purpose of remineralization.Therefore,the purpose of this study was to prepare carboxymethyl chitosan（CMC）and lysozyme（LYZ）nanogels in order to achieve the stability and controlled-delivery of amorphous calcium phosphate（ACP）which was dental hard tissue mineralization precursor,to observe whether mineral layers could be formed on the surface of demineralized enamel and dentin,which provide a novel idea and method for the treatment of enamel demineralization and dentin hypersensitivity.Methods1.CMC/LYZ-ACP nanogels loaded with amorphous calcium phosphate（ACP）were prepared by adding potassium dihydrogen phosphate（KH₂PO₄）and calcium chloride dihydrate（Ca Cl₂×2H₂O）into CMC and LYZ nanogels with different proportions.The morphologies of the nanogels were measured by transmission electron microscope（TEM）.The size and zeta potential of the nanogels were measured by dynamic light scattering/particle electrophoresis,in order to explore the optimal ratio of CMC and LYZ for electrostatic assembly during the preparation of nanogels.2.Different concentrations of sodium chloride（Na Cl）and different p H values were used to explore the optimal conditions for the dissociation of CMC/LYZ-ACP nanogels to release ACP.The morphology,particle size and zeta potential of the samples were determined by TEM and dynamic light scattering/particle electrophoresis.UV absorption and intrinsic fluorescence were used to further prove the effect of Na Cl and p H value on CMC/LYZ-ACP nanogels.3.CMC/LYZ-ACP nanogels were applied to the demineralized enamel surface.The remineralization effect of CMC/LYZ-ACP nanogels on demineralized enamel surface was detected by the nano-indentation test,X-ray diffraction（XRD）,confocal laser scanning microscope（CLSM）,and scanning electron microscope（SEM）,compared with the fluoride treatment group.The same methods were applied to observe the sealing effect of dentinal tubules of CMC/LYZ-ACP nanogels on the dentinal tubules surface,compared with Green Or^TMdesensitizer.4.The remineralized enamel and dentin blocks were fixed into the rat mouth which was simulated human oral environment by saliva scouring and chewing friction in the rat oral cavity.SEM and CLSM were used to detect the effects of oral environment on the surface morphology and permeability of the samples.Results1.The optimum ratio of CMC and lysozyme was 10:8.CMC/LYZ-ACP nanogels were spherical structures with negative charge and particle size of approximately 300 nm.2.In the presence of 0.15 M Na Cl,ACP was released from the disassembled nanogels and transformed into hydroxyapatite（HAP）at p H 8.3.The group treated with CMC/LYZ-ACP nanogels showed the regeneration of an aprismatic enamel-like layer on demineralized enamel surface which was higher mechanical properties（P<0.05）compared to those of the fluoride-treated group.Also,mineralized layer could be formed on the exposed dentinal tubules surface,and occluded the interior of tubules,which had a high impermeability（P<0.01）compared with the Green Or^TMdesensitizer.4.SEM showed that a dense mineral layer still covered the surface of dental hard tissue treated with CMC/LYZ-ACP nanogels,and CLSM results confirmed its impermeability in oral environment.ConclusionsCMC/LYZ-ACP nanogels can encapsulate ACP and control its delivery,forming a dense mineral layer on the surface of demineralized dental hard tissue,which can still achieve good stability and sealing effect in rat oral cavity.It has a certain application prospect in the prevention and treatment of enamel demineralization and dentin hypersensitivity.