Recently, the term tensotaxis was proposed to describe the phenomenon that tensile stress or strain affects cell migration. Even so, less attention has been paid to the effects of compressive stress on cell behavior. In this study, by using an injection-molded method combined with photoelastic technology, we developed residual stress gradient-controlled poly-L-lactide discs. After culturing NIH-3T3 fibroblasts on the stress gradient substrate, the cell distributions for high- and low-stress regions were measured and compared. Our results showed that there were significantly more cells in the low-compressive stress region relative to their high-stress analogs (p < 0.05). In addition, NIH-3T3 fibroblasts in the low-compressive stress region expressed more abundant extensive filopodia. These findings provide greater insight into the interaction between cells and substrates, and could serve as a useful reference for connective tissue development and repair.