The physiochemical characteristics of a material with in vivo applications are critical for the clinical success of the implant and regulate both cellular adhesion and differentiated cellular function. Topographical modification of an orthopaedic implant may be a viable method to guide tissue integration and has been shown in vitro to dramatically influence osteogenesis, inhibit bone resorption and regulate integrin mediated cell adhesion. Integrins function as force dependant mechanotransducers, acting via the actin cytoskeleton to translate tension applied at the tissue level to changes in cellular function via intricate signalling pathways. In particular the ERK/MAPK signalling cascade is a known regulator of osteospecific differentiation and function. Here we investigate the effects of nanoscale pits and grooves on focal adhesion formation in human osteoblasts (HOBs) and the ERK/MAPK signalling pathway in mesenchymal populations. Nanopit arrays disrupted adhesion formation and cellular spreading in HOBs and impaired osteospecific differentiation in skeletal stem cells. HOBs cultured on 10 microm wide groove/ridge arrays formed significantly less focal adhesions than cells cultured on planar substrates and displayed negligible differentiation along the osteospecific lineage, undergoing up-regulations in the expression of adipospecific genes. Conversely, osteospecific function was correlated to increased integrin mediated adhesion formation and cellular spreading as noted in HOBS cultured on 100 microm wide groove arrays. Here osteospecific differentiation and function was linked to focal adhesion growth and FAK mediated activation of the ERK/MAPK signalling pathway in mesenchymal populations.