Engineering osteoinductive hydroxyapatite via zinc and strontium co-substitution

Critical-size bone defects remain a major clinical challenge due to the limited regenerative capacity of bone tissue, necessitating biomaterials that provide both structural support and biological stimulation. Synthetic hydroxyapatite (HAp) is widely used for bone repair because of its close similarity to native bone minerals; however, strategies to enhance its biological performance are actively being explored. Among these, ionic substitution-particularly with zinc (Zn²⁺) and strontium (Sr²⁺) ions-has emerged as a promising approach to promote osteogenesis while inhibiting bone resorption. Recent studies on Zn/Sr-substituted HAp have reported the formation of undesirable secondary phases, which compromise biological performance. Considering these limitations, the present study aims to synthesize phase-pure Zn- and Sr-substituted hydroxyapatite nanoparticles using a soft-template method, with careful optimization of Zn²⁺ and Sr²⁺ concentrations for bone tissue engineering applications. Accordingly, Zn-substituted, Sr-substituted, and Zn/Sr co-substituted hydroxyapatite nanoparticles (Zn-HAp, Sr-HAp, and ZnSr-HAp) were synthesised and systematically characterised. Physicochemical analysis revealed that, in comparison with single-ion substitution, Zn/Sr co-substitution significantly altered crystal nucleation and growth behaviour while preserving phase purity. Biological evaluation using bone-derived cells demonstrated that ZnSr-HAp nanoparticles were highly biocompatible and significantly enhanced cell attachment, proliferation, and matrix mineralisation compared to pristine and single ion-substituted HAp. Furthermore, ZnSr-HAp treatment markedly upregulated the expression of collagen type I (COL I) and osteocalcin (OCN), indicating the promotion of both early extracellular matrix formation and late-stage osteogenic maturation. Collectively, these findings demonstrate a synergistic effect of Zn and Sr co-substitution in enhancing the osteoinductive and osteoconductive properties of HAp. Overall, Zn/Sr co-substituted hydroxyapatite nanoparticles represent a promising bioactive platform for advanced bone regeneration applications.