Stem cells reside within instructive and tissue-specific niches in the body, such as complex and controlled biochemical mixtures of soluble and insoluble factors. In particular, it is widely accepted that stem cells display high sensitivity to the extracellular matrix (ECM) composed of complex and well-defined nanostructures of protein fibers, suggesting that the use of nanotopographical cues hold great potentials to control stem cell functions. The structure of the natural ECM in various tissues including bone, tooth, nerve, and heart usually reveals highly oriented grooved structures with various length scales in nanometers. Inspired by such ultrastructural observations, we designed and fabricated nanogrooved matrices with various groove densities using ultraviolet (UV)-assisted capillary force lithography (CFL). It is hypothesized that the defined nanotopographical density would provide unique topographical cues to control the morphology and function of stem cells.