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Background Previous studies have demonstrated increased functions of osteoblasts on nanophase materials compared to conventional ceramics or composites.Nanophase materials are unique materials that simulate dimensions of constituent components of bone as they possess particle or grain sizes less than 100 nm.However,to date,interactions of osteoblasts on nanophase materials compared to conventional metals remain to be elucidated.The objective of the present in vitro study was to synthesize nanophase metals (Ti6Al4V),characterize,and evaluate osteoblast functions on Ti6Al4V.Such metals in conventional form are widely used in orthopedic applications.Methods In this work,nanophase Ti6Al4V surfaces were processed by the severe plastic deformation (SPD) principle and used to investigate osteoblast long-term functions.Primary cultured osteoblasts from neonatal rat calvaria were cultured on both nanophase and conventional Ti6Al4V substrates.Cell proliferation,total protein content,and alkaline phosphatase (ALP) activity were evaluated after 1,3,7,10 and 14 days.Calcium deposition,gene expression of type Ⅰ collagen (Col-Ⅰ),osteocalcin (OC),osteopontin (OP) and the production of insulin-like growth factor-Ⅰ (IGF-Ⅰ) and transforming growth factor-beta 1 (TGF-β1) were also investigated after 14 days of culture.Results Functions of osteoblasts including proliferation,synthesis of protein,and ALP activity were improved on the nanophase compared to the conventional Ti6Al4V.The expression of Col-Ⅰ,OC and OP mRNA was also increased on nanophase Ti6Al4V after 14 days of culture.Calcium deposition was the same;the average number of the calcified nodules on the two Ti6Al4V surfaces was similar after 14 days of culture;however,highly significant size calcified nodules on the nanophase Ti6Al4V was observed,Of the growth factors examined,only TGF-β1 showed a difference in production on the nanophase surface.Conclusion Nanophase Ti6Al4V surfaces improve proliferation,differentiation and mineralization of osteoblast cells and should be further considered for orthopedic implant applications.