Abstract
Biomimetic synthesis of artificial enamel is a promising strategy for the prevention and restoration of defective enamel. We have recently reported that a hydrogel system composed of chitosan-amelogenin (CS-AMEL) and calcium phosphate is effective in forming an enamel-like layer that has a seamless interface with natural tooth surfaces. Here, to improve the mechanical system function and to facilitate the biomimetic enamel regrowth, matrix metalloproteinase-20 (MMP-20) was introduced into the CS-AMEL hydrogel. Inspired by our recent finding that MMP-20 prevents protein occlusion inside enamel crystals, we hypothesized that addition of MMP-20 to CS-AMEL hydrogel could reinforce the newly grown layer. Recombinant human MMP-20 was added to the CS-AMEL hydrogel to cleave full-length amelogenin during the growth of enamel-like crystals on an etched enamel surface. The MMP-20 proteolysis of amelogenin was studied, and the morphology, composition, and mechanical properties of the newly grown layer were characterized. We found that amelogenin was gradually degraded by MMP-20 in the presence of chitosan. The newly grown crystals in the sample treated with MMP-20-CS-AMEL hydrogel showed more uniform orientation and greater crystallinity than the samples treated with CS-AMEL hydrogel without MMP-20. Stepwise processing of amelogenin by MMP-20 in the CS-AMEL hydrogel prevented undesirable protein occlusion within the newly formed crystals. As a result, both the modulus and hardness of the repaired enamel were significantly increased (1.8- and 2.4-fold, respectively) by the MMP-20-CS-AMEL hydrogel. Although future work is needed to further incorporate other enamel matrix proteins into the system, this study brings us one step closer to biomimetic enamel regrowth.