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Modified layers with multiscale structures have been extensively explored and rationally designed on Ti-based orthopedic implants recently, since the remarkable improvement of biomedical properties. In consideration of the inefficiency, costliness and waste products of existing technologies, herein, we proposed the ultrasonic shot peening and induction heating protocol to construct the micro-nano complex oxide structures on Ti6Al4V in term of its advantages of environmental friend, rapid heating, low-cost and easy to process. With the USP pretreatment and induction heating, the nonuniform oxide crystallites firstly appeared on smooth surface and successfully grow up into the well-distributed TiO 2 nanocrystallites, and transformed to netlike structures with submicro-sized worms when subjected to high-power of shot peening, which was significantly influenced by the microstructural evolution of alloy matrix. The submicro/nano-structured oxide layers mainly contained rutile/anatase TiO 2 and little Al 2 O 3 between grain boundaries, successively enhanced the surface roughness and wettability, and tended to form the hydrophilic surfaces. The adhesion strength of the oxide layers to substrates and hardness were also distinctly increased with increasing the IHT period from 20 to 35 s. In vitro BMSCs culturing proved that the prepared submicro/nano-scale oxide layers possessed favorable cell adhesion and proliferation, and further improved the osteogenic differentiation. The novelty of this work was to explored the evolution law of submicro-and nano-scale structures in osteogenic differentiation, and further better clinical application of Ti-based biomaterials. Image 1 • Ultrasonic shot peening and induction heating treatment were jointly proposed. • Microstructure evolution of alloy matrices promoted structure transformation of oxide layers. • Surface roughness, wettability and hardness were markedly improved. • Nano/submicro-structured oxide layers possessed good stem cell compatibility. • Oxide layers with multi-structures further promoted osteogenic differentiation of BMSCs. [ABSTRACT FROM AUTHOR]