A vascular cell culture patterning substrate, which can efficiently form a plurality of blood vessels on one substrate. The vascular cell culture patterning substrate includes: a base material; a vascular cell adhesion portion formed in at least two substantially parallel lines on the base material, and having adhesive properties to a vascular cell which forms a blood vessel; and a vascular cell adhesion-inhibiting portion formed in between two adjacent vascular cell adhesion portions on the base material, and inhibiting adhesion to the vascular cell. The vascular cell adhesion-inhibiting portion contains a vascular cell adhesion-inhibiting material having vascular cell adhesion-inhibiting properties of inhibiting adhesion to the vascular cell.