The environment around a cell during in vitro culture is unlikely to mimic those in vivo. Preliminary experiments with nanotopography have shown that nanoscale features can strongly influence cell morphology, adhesion, proliferation and gene regulation, but the mechanisms mediating this cell response remain unclear. In this study a well defined nanotopography, consisting of 100 nm wide and 160 nm high cylindrical columns, was used in fibroblast culture. In order to build on previously published morphological data that showed changes in cell spreading on the nanocolumns, in this study gene regulation was monitored using a 1718 gene microarray. Transmission electron microscopy, fluorescent observation of actin and Rac and area quantification have been used to re-affirm the microarray observations. The results indicate that changes in cell spreading correlate with a number of gene up- and down-regulations as will be described within the manuscript.