Stem cell response to a library of scaffolds with varied 3D structures was investigated. Microarray screening revealed that each type of scaffold structure induced a unique gene expression signature in primary human bone marrow stromal cells (hBMSCs). Hierarchical cluster analysis showed that treatments sorted by scaffold structure and not by polymer chemistry suggesting that scaffold structure was more influential than scaffold composition. Further, the effects of scaffold structure on hBMSC function were mediated by cell shape. Of all the scaffolds tested, only scaffolds with a nanofibrous morphology were able to drive the hBMSCs down an osteogenic lineage in the absence of osteogenic supplements. Nanofiber scaffolds forced the hBMSCs to assume an elongated, highly branched morphology. This same morphology was seen in osteogenic controls where hBMSCs were cultured on flat polymer films in the presence of osteogenic supplements (OS). In contrast, hBMSCs cultured on flat polymer films in the absence of OS assumed a more rounded and less-branched morphology. These results indicate that cells are more sensitive to scaffold structure than previously appreciated and suggest that scaffold efficacy can be optimized by tailoring the scaffold structure to force cells into morphologies that direct them to differentiate down the desired lineage.