AbstractThis study explores the enhancement of biocompatible titanium‐based implants through surface functionalization for improved bone healing. Specifically, a near‐beta type Ti‐13Nb‐13Zr alloy is 3D printed using laser powder bed fusion and subsequently textured using nanosecond (ns) and picosecond (ps) direct laser interference patterning (DLIP) to create single‐scale and multi‐scale surface textures. On these textures, the cell behavior, morphology, metabolic activity and osteogenic differentiation potential of human bone marrow stromal cells are assessed using fluorescence microscopy and MTS assays. Moreover, tissue non‐specific alkaline phosphatase activity served as an early osteoblast production marker. Compared to untextured specimens, both types of textures exhibited higher metabolic activity and cell proliferation. Single‐scale ns‐DLIP textures encouraged cell extensions anchored in groove regions, while ps‐DLIP textures with hierarchical structures promoted cell extensions attaching to nanostructures on sidewalls. The groove width and nanotopographies in groove areas facilitated cell spreading. Surface topography, roughness, and surface chemistry (surface energy, wettability) influenced cell adhesion, proliferation, and differentiation. A comprehensive evaluation of DLIP‐generated surface textures, including their topography and chemical states, complements the factors affecting in vitro cell behavior. Overall, this research demonstrates the potential of surface‐functionalized 3Dprinted titanium for a novel generation of biocompatible implants.