•The Ni-Ti-O nanotubes (NTs) with similar surface properties but different Ni release levels are fabricated on NiTi alloy.•The NTs release more Ni than NiTi control but their cytocompatibility is comparable.•The NTs possess different Ni release levels but possess similar cytocompatibility. To elucidate the relationship between Ni release and cytocompatibility of NiTi alloy, Ni-Ti-O nanotubes (NTs) are produced on its surface by anodization followed by annealing. The NiTi control has higher corrosion current density but releases less Ni than anodized samples, suggesting chemical dissolution of the NTs also contributes to Ni release. Although the annealed NTs release more Ni than that of NiTi control, they possess similar cytocompatibility. In addition, cytocompatibilitiy of the NTs with similar surface properties but have different Ni release levels is also comparable. These results suggest the amount of released Ni has little influence on the cytocompatibility.