Mesenchymal stem cells (MSCs) with the ability to differentiate into various mesoderm-like cells are known to migrate to various organs to repair injured tissues. They can attach to the implant surface, differentiate into bone-forming cells, and ultimately osseointegrate with the prosthesis. This study investigates bone marrow-derived mesenchymal stem cellular response to the grain structure of titanium substrates produced by high-pressure torsion and annealing processes. Cell attachment, proliferation, viability, and morphology are evaluated on the surface of differently processed nanostructured and coarse-grained samples. The bacterial adhesion and calcium phosphate crystal formation and growth are also assessed on the surface of the substrates. The nanostructured titanium shows significantly higher cell adhesion, proliferation, spreading, and viability compared with the untreated and coarse-grained titanium substrates. The adhesion of bacteria is lower and surface bioactivity is higher on the surface of the nanostructured titanium substrate. The results demonstrate the superior MSC compatibility, antibacterial efficacy, and surface bioactivity of the nanostructured titanium substrates, which could lead to early implant fixation and improved osseointegration. Copyright © 2012 John Wiley & Sons, Ltd.