Cellular behavior on TiO2 nanonodular structures in a micro-to-nanoscale hierarchy model

Biological tissues involve hierarchical organizations of structures and components. We created a micropit-and-nanonodule hybrid topography of TiO2 by applying a recently reported nanonodular self-assembly technique on acid-etch-created micropit titanium surfaces. The size of the nanonodules was controllable by changing the assembly time. The created micro-nano-hybrid surface rendered a greater surface area and roughness, and extensive geographical undercut on the existing micropit surface and resembled the surface morphology of biomineralized matrices. Rat bone marrow-derived osteoblasts were cultured on titanium disks with either micropits alone, micropits with 100-nm nodules, micropits with 300-nm nodules, or micropits with 500-nm nodules. The addition of nanonodules to micropits selectively promoted osteoblast but not fibroblast function. Unlike the reported advantages of microfeatures that promote osteoblast differentiation but inhibit its proliferation, micro-nano-hybrid topography substantially enhanced both. We also demonstrated that these biological effects were most pronounced when the nanonodules were tailored to a diameter of 300 nm within the micropits. An implant biomechanical test in a rat femur model revealed that the strength of bone–titanium integration was more than three times greater for the implants with micropits and 300-nm nanonodules than the implants with micropits alone. These results suggest the establishment of functionalized nano-in-microtitanium surfaces for improved osteoconductivity, and may provide a biomimetic micro-to-nanoscale hierarchical model to study the nanofeatures of biomaterials.