Immobilization of phosphate monomers on collagen induces biomimetic mineralization

• Published in: Bio-medical materials and engineering 2015-01, Vol.25 (1), p.89-99
• Main Authors: Nurrohman, Hamid, Nakashima, Syozi, Takagaki, Tomohiro, Sadr, Alireza, Nikaido, Toru, Asakawa, Yuya, Uo, Motohiro, Marshall, Sally J., Tagami, Junji
• Format: Article
• Language: English
• Subjects: Animals; Apatites - chemistry; Binding; Biocompatible Materials - chemistry; Biomimetics; Cattle; Collagen - chemistry; Collagen Type I - chemistry; Collagens; Crystallization; Crystallography, X-Ray; Esters - chemistry; Ethyl alcohol; Immobilization; Methacrylates - chemistry; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Mineralization; Minerals; Monomers; Phosphates; Phosphates - chemistry; Phosphoproteins - chemistry; Phosphorylation; Protein Binding; Spectroscopy, Fourier Transform Infrared; Temperature; Transmission electron microscopy
• ISSN: 0959-2989; 1878-3619
• DOI: 10.3233/BME-141243

BACKGROUND: Immobilization of phosphoproteins on type-I collagen via covalent binding may induce extra- and intrafibrillar mineralization. OBJECTIVE: This study tested the hypothesis that methacrylate phosphate esters immobilized on reconstituted type-I collagen can mimic the nucleating role of phosphoproteins. METHODS: Three functional monomers (MDP, GPDM and Phenyl-P) that differed in chemical structure and steric hindrances around the phosphate moiety were evaluated. Reconstituted type-I collagen was either left untouched (control) or treated by 5% monomer/ethanol for 20 s. All samples were incubated in simulated dentinal fluid as mineralizing medium at 37°C for 7 or 14 days. The extra- and intrafibrillar mineralization were examined by SEM and TEM/SAED crystallography, respectively. RESULTS: FT-IR spectroscopy showed that the phosphate groups were incorporated on reconstituted collagen, irrespective of their chemical structure. MDP immobilization induced dense growth of extrafibrillar mineral over time, while with GPDM- and Phenyl-P-immobilized collagen, mineralization was moderate and sparse, respectively. TEM/SAED evidence disclosed that intrafibrillar minerals exclusively occurred in MDP-immobilized collagen. CONCLUSIONS: Immobilization of MDP, which had the lowest steric hindrance, could induce significant biomimetic extra- and intrafibrillar mineralization; resembling the lowest level of hierarchy organization of dentin.