Hydroxyapatite (HA) is a bioceramic very similar to the mineral component of bones and teeth. It is well established that osteoblasts grow better onto HA-coated metals than on metals alone. Herein, the preparation of a new system consisting of magnetite (Fe3O4) and HA functionalized with oleic acid and simvastatin (SIMV), and incorporated in chitosan (CHI) scaffolds, was undertaken. HA was synthesized by the hydrothermal method, while Fe3O4 was synthesized by co-precipitation. The polymer matrix was obtained using a 2% CHI solution, and allowed to stir for 2 h. The final material was freeze-dried to produce scaffolds. The magnetic properties remained unchanged after the formation of the composite, as well as after the preparation of the scaffolds, maintaining the superparamagnetism. CHI scaffolds were analyzed by scanning electronic spectroscopy (SEM) and showed a high porosity, with very evident cavities, which provides the functionality of bone growth support during the remineralization process in possible regions affected by bone tissue losses. The synthesized composite showed an average particle size between 15 and 23 nm for particles (HA and Fe3O4). The scaffolds showed considerable porosity, which is important for the performance of various functions of the tissue structure. Moreover, the addition of simvastatin in the system can promote bone formation.
Objectives
We aimed to evaluate the effect of nanohydroxyapatite morphology and its interaction with anionic collagen on osteoblast activity.
Materials and Methods
Murine osteoblasts were incubated with a commercial collagen scaffold (as a control) or collagen-nanohydroxyapatite scaffolds (Col-HANP) for 24 and 48 hours for viability and proliferation assessments by MTT and Ki67 immunofluorescence, respectively. The hydroxyapatite nanoparticles were synthesized in three different morphologies/sizes (labeled as Col-HANP 0h, as Col-HANP 2h, and as Col-HANP 5h) as a function of the hydrothermal synthetic approach. Osteoblast's activity was investigated by bone alkaline phosphatase activity (ALP) and Von Kossa mineralization assays. For biocompatibility evaluation, the scaffolds were implanted subcutaneously in the dorsum of male Wistar rats for 7 and 15 days.
Results
The incubation of cells with Col-HANP 5h for 48h resulted in a significant increase in their proliferation and activity. The implantation of Col-HANP 5h in the subcutaneous tissue presented decreased recruitment of inflammatory cells and IL-1β levels on day 7, as well as an increase in collagen synthesis on day 15 compared to collagen and control groups.
Conclusions
The significant effects on osteoblasts proliferation and activity illustrate the potential application of Col-HANP 5h scaffold as a promising strategy for bone tissue engineering.
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