Hierarchical nanowire-constructed TiO2 microspheres (NW-TMS) were successfully synthesized from TiO2 hollow microspheres (THMS) via an in situ transformation in hydrothermal condition followed by an annealing process in air, while the hydrogenated nanowire-constructed TiO2 microspheres (HNW-TMS) were obtained after annealing in hydrogen. The possible formation mechanisms of HNW-TMS have been proposed. After morphology transformation, the specific surface area of hierarchical nanomaterials increases significantly, and the network structure formed by nanowires improves the structure stability of NW-TMS to tolerate the volume change during lithium-ion intercalation/extraction. In fact, NW-TMS and HNW-TMS exhibit much improved performance of Li-ion batteries (LIBs) than original hollow TiO2 microspheres. Especially, hydrogenated nanowire-constructed TiO2 microspheres demonstrate a high capacity of 284.5 mA h g−1 at 50 mA g−1 (253.0 and 223.6 for NW-TMS and THMS), excellent rate capacity of 141.1 mA h g−1 at 1000 mA g−1 after 200 cycles (114.5 and 74.1 for NW-TMS and THMS), and good cyclic stability (82.8% capacity retention after 200 cycles at 1000 mA g−1). The enhanced LIB performance can be attributed to the reduced particle size and increased surface area of active materials, 3D network-like microstructures formed by nanowires, and more active sites induced by more oxygen vacancies in the HNW-TMS.