Surface-induced thrombosis and lack of endothelialization are major drawbacks that hamper the widespread application of polyurethanes for the fabrication of implantable cardiovascular devices. Endothelialization of the blood-contacting surfaces of these devices may avoid thrombus formation and may be implemented by strategies that introduce micro and submicron patterns that favor adhesion and growth of endothelial cells. In this study, we used laser radiation to directly introduce topographical patterns in the low micrometer range on castor oil-based polyurethane, which is currently employed to fabricate cardiovascular devices. We have investigated cell adhesion, proliferation, morphology and alignment in response to these topographies. Reported results show that line-like and pillar-like patterns improved adhesion and proliferation rate of cultured endothelial cells. The line-like pattern with 1 μm groove periodicity was the most efficient to enhance cell adhesion and induced marked polarization and alignment. Our study suggests the viability of using laser radiation to functionalize PU-based implants by the introduction of specific microtopography to facilitate the development of a functional endothelium on target surfaces.