Influence of Ca/P in Modified-Simulated Body Fluid Solution on Intrafibrillar Mineralization of Collagen with Polydopamine to Generate Collagen-PDA-HAp Scaffolds

Large bone defects have attracted attention as a remain major challenge in bone tissue engineering. In bone tissue, the mineralized collagen fibril is an important basic building block which collagen molecules are hierarchically assembled into fibrils and mineralized with the formation of hydroxyapatite (HAp) nanocrystals. Due to the uniformly distribution HAp in collagen fibrils and together built up hierarchically from nano to macrostructures in bone tissue, mineralized collagen fibril is regards as the important level for the mechanical properties. Collagen based scaffolds have been increasing used in tissue engineering due to its excellent biocompatibility. However, the insufficient mechanical strength and structural support limit it wider in bone regeneration application. Many attempts have been devoted to improve collagen-HAp based scaffolds’ properties by understanding the process of bone mineralization to fabricate bone mimicking scaffolds. The mineralization pathway of HAp has been suggested as non-classical crystallization as reported, but the physical and chemical mechanism is still poorly understood with a difficult challenge, especially in collagen fibril’s structure. In this work, with the presence of polydopamine (PDA) to promote the biomineralization process, the structure of intrafibrillar mineralized collagen-PDA fibrils in modified-simulated body fluid (m-SBF) solution was studied at the nano scale by STEM-EDS techniques to investigate a detail hydroxyapatite intrafibrillar mineralization pathway. Collagen-amorphous calcium phosphate (ACP) fibrils was obtained by assembling Collagen-PDA fibrils in SBF solution with the present of Mg²⁺ and polyaspartic acid as a precursor stabilizer. Then later the phase transformation of ACP to HAp was determined by tuning the phosphate concentrations in m-SBF. It was found, in this work, that the phase transformation of ACP to HAp in Collagen fibrils can be accelerated even in 12 hr-mineralization with the m-SBF solution of calcium and phosphate ratio of 1:10, higher phosphate ratio produces a monodentate Ca-P geometry and evolved directly to HAp, while the lower 1:5 would affect in slower phase transformation kinetic, and for 1:1 the crystallization of HAp cannot be initiated even up to 7 days of mineralization. The finding suggests that an elevated concentration of phosphate is crucial for initiate phase transformation of ACP to HAp. From these studies and strategy, we adopted to fabricate mechanically self-sustained Collagen-PDA-HAp hydrogels. By controlling the mineralization, it is beneficial for collagen-based scaffolds’ fabrication design. A synergistic effect was also investigated and observed in the Collagen-PDA-ACP and Collagen-PDA-HAp for both in vitro osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). Thus, the results presented in this work could serve a better understanding the mechanism of the HAp mineralization in collagen fibrils, which may provide approaches to effective, and materials design for efficient osteogenesis bone tissue engineering.