Repair Of Demineralized Dentin By Using Nano-amorphous Calcium And Its Effects On Structure And Mechanical Behavior Of The Dentin

As demonstrated by a number of recent studies, bone and teeth are typical biomineral tissues, formed through a series of transformation and crystallization of amorphous calcium phosphate (ACP) under the control of biological macromolecules in organism. As precursor mineral phase and being of isotropic and liquidity, ACP infiltrates into the collagen matrix resulting in organic-inorganic complex, which has excellent mechanical properties. Nevertheless, ACP phase is quite unstable under pure aqueous, tending to evolve into the more thermodynamic stable phase HAP and losing the feature of isotropic and liquidity. At present, many scholars attempt to repair the demineralized dentin by using the Biomimetic Mineralization Perspective. The mainstream studies are focusing on combining two kinds of non-collagen analogues with a solution containing calcium and phosphate ion source. This method has formed intrafibrillar mineral in the collagen matrix, which is the standard for biomimetic collagen fibrils mineralization. However, remineralization in this system took nearly4months, thus significantly limits its clinical application prospects. Here we conducted remineralization of the demineralized dentin under various content PAA-ACP stabilizer, in order to solve this problem.Objective:There are three objections for this study. First we explored the relationship between the PAA content and stabilization time of ACP. The second is to inspect the process of remineralization by conducting dentin remineralization in the three remineralization mediums. At last, both the nanomechanical and the nanostructure of the remineralized were characterized and the relationship between them was researched.Methods:1. PAA of three concentrations,100μg/ml,500μg/ml and1000μg/ml, were chosen in the study after the previous experiments. The phase transformation of ACP under the three concentrations was traced with FTIR, morphology of the precipitates was characterized by TEM. 2. The third molars were used in this study. Dentin slides of1mm thickness were obtained by cutting perpendicularly to the long axis of tooth under water cooling. Before remineralization, the slides were demineralized with the35%phosphoric acid and followed by abundant water rushing. The30dentin slides were divided into three groups (N=10), placing in the three remineralization mediums respectively. Then the mediums with dentin slides in were cultured in37.5℃oven. The result showed that dentin was perfectly remineralized only in the medium with500μg/m PAA. Thus dentin sample in the medium were extracted at day1,3,5,7, respectively, and the mineralization process was tracked by using SEM and TEM.3.2dentin slides were picked out from the solution with500μg/ml PAA on the fifteenth day when remineralization is completed. One of the slides was embedded with epoxy resin, cut into80nm thick slices and then being observed by using TEM. The other was one was tested by nanomdentation with its surface being kept wet.Results:1. The transformation time from ACP to HAP is in direct proportion with the PAA concentration. Under the three concentrated PAA, transformation time needed are approximately8h,-15d,>30d, respectively.2. The dentin remineralization effect is highly related to PAA basing on its concentration. Among the three PAA concentrations, the dentin were perfectly remineralized in the medium with500μg/ml PAA, whereas PAA of low concentration being not able to stabilize ACP and high content PAA significantly inhibits the phase transformation of ACP, both lead to failure of dentin remineralization.3. The process of dentin remineralization was tracked by observing the remineralized dentin at different time points using SEM and TEM. The result shows that the process of dentin remineralization is not a one-step process but is achieved through gradually phase transformation from ACP to HAP. 4. After15days of remineralization, the super structure of the remineralized dentin was characterized by HRTEM and the mechanical behavior of the dentin was tested by using nanoindentation. The results show that both the ultrastructure and the mechanical behavior were restored after the remineralization.Conclusion:The concentration of PAA is an important parameter both for ACP crystallization and dentin remineralization. The stabilization time of ACP is in direct proportion with the PAA content. By adjusting concentration of PAA, we can control the liquidity and penetration ability of ACP, which will further its effect on dentin remineralization. In the presence of500μg/1PAA, demineralized dentin was perfectly remineralized and the resulting dentin is of excellent mechanical behavior. In addition, the process of dentin remineralization is not a one-step process but is achieved through gradually phase transformation from ACP to HAP. Our study may provide promising application for hard tissues regeneration in the future.