Constructing hydroxyapatite/collagen nanocomposite scaffold via in-situ mineralization and comparing in-situ scaffolds and mix-powder in bone defect regeneration in rabbit animal model

Aim and Background: In situ mineralization is a simple and effective technique that allows for the fabrication of homogeneous composites with unique morphology and well-distributed HA in hydrogels. In situ synthesis of HA particles in the presence of collagen fibers is a biomimetic approach that can lead to formation of collagen/HA composites similar to bone. This study aimed to compare powder-mixed HA/Co hydrogel and in situ synthesized HA/Co hydrogel on the regeneration of bone defect in the rabbit animal model. Methods: An in-situ collagen/HA scaffold was synthesized by adding HA salts to a collagen acidic solution to mineralize HA in hydrogel at physiological conditions (pH = 7.4 and 37 °C). Under the same conditions, a mix-powder collagen/HA scaffold was prepared, but instead of HA salts, HA powder was added. Following that, physicochemical properties were investigated. Then, in-vitro tests were performed, including cytocompatibility assays, alkaline phosphatase activity assays, and alizarin red staining. Afterwards, in-vivo analysis was performed on a critical-sized bone defect in a rabbit animal model, followed by radiological evaluation and microscopical examination. Results and discussion: The plate-like HA particles are distributed homogeneously in the in situ HA/Co scaffold compared to powder-mixed HA/Co scaffold and had a similar structure to bone with carbonated plate-like HA particles and nanofibrilated Co matrix. These properties along with excellent cytocompatibility and osteogenicity of in situ HA/collagen nanocomposite candidate it as a proper scaffold for bone tissue engineering. Conclusion: In situ HA/Co nanocomposite can accelerate bone regeneration due to osteoblastic production of osteocalcin protein, making it a new approach for bone tissue engineering