Micro/nano hierarchical structures and nano-sized hydroxyapatite (nHA) have been proven to play a critical role in promoting osseointegration. However, still, it is a challenge to prepare tightly adherent nHA coatings with micro/nano hierarchical structure, and the correlation between the physicochemical properties of hydroxyapatite (HA) and osteoblasts (OBs) activity at different stages is unclear yet. In this study, the grain refinement of large-sized HA generated by traditional microarc oxidation-hydrothermal treatment was achieved through a simple method of adding Ca/P (Ca2+ and PO43−) to hydrothermal reaction, and micro/nano hierarchical nHA coatings with different crystallinities and grain sizes were prepared on Zr16Nb12Ti alloy. Further, the OBs responses on these coatings were investigated in proliferation and differentiation stages, respectively. It was found that small-sized HA particles provide more integrin-binding sites, which are important for the adhesion and proliferation stages of OBs. The high ion release resulting from the low crystallinity of HA is a crucial condition for promoting OBs differentiation. However, amorphous HA coatings with excessively low crystallinity exhibit poor osteogenic activity due to their inferior cell adhesion activity. Finally, a micro/nano hierarchical nHA coating with appropriate crystallinity was prepared through a simple method, demonstrating excellent osteogenic activity, and providing new strategies for biomimetic coatings research.
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