Tailorable hierarchical structures of biomimetic hydroxyapatite micro/nano particles promoting endocytosis and osteogenic differentiation of stem cells
Hydroxyapatite (HA) micro/nano particles have great promise for artificial bone and dental substitutes, or drug carrier systems. However, the precise regulation of hydroxyapatite micro/nano particles with controllable physicochemical properties（as hierarchical structure, particle size, potential and crystallinity） is still a challenge. And the further effects of different hierarchical structures on biological responses have been rarely reported. Herein, hydroxyapatite particles with precisely tailored micro/nano hierarchical structure have been developed with an elaborate biomimetic synthesis technology. Three representative microspheres, namely, micro/nano needle-like HA microspheres, micro/nano rod-like HA microspheres, and micro/nano flake-like HA microspheres, were featured to evaluate their biological responses with stem cells. The pore structure facilitated the adsorption of serum adhesive proteins, which together with the unique hierarchical architecture of micro/nano flake-like HA microspheres remarkably promoted the endocytosis efficiency in a concentration-dependent manner. The qRT - PCR together with RNA-seq and Western blot tests showed that micro/nano flake-like HA microspheres more significantly up-regulated the genes expression and proteins production related to osteogenic differentiation among three microspheres through the activated ERK/MAPK signaling. RNA-seq further revealed a complex mechanism of cell interface events, suggesting that hierarchical architecture of HA microspheres were of crucial importance for the regulation of actin cytoskeleton involved in the modulation of cell adhesion which positively stimulated osteogenic differentiation of stem cells. Moreover, the endocytosis of microspheres into lysosomes brought about an increase in intracellular Ca2+ levels, which activated a possible intracellular Ca2+ - mediated signaling cascades (Ras / cAMP / Rap1 / MAPK signal pathways) related to osteogenic differentiation of stem cells. Our findings shed light on the effects of the different hierarchical structure of HA microspheres on stem cell differentiation and contribute to the optimal design of implant materials.