Establishing aligned microvessel-like structures is crucial to the engineered construct in vitro for the efficient tissue perfusion. In this study, human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells derived from induced pluripotent stem cells (iPSC-MSCs) were co-cultured on aligned and random scaffolds to verify the hypothesis that contact guidance could direct the formation of aligned microvessel-like structures. As tested, the major discrepancy between aligned and random scaffolds lied in the fiber orientation. Investigated by immunofluorescence staining, co-culturing HUVECs and iPSC-MSCs could form the microvessel-like structures. Applying this co-culturing system to both scaffolds showed that the orientation of microvessel-like structures was closely related to the fiber orientation of scaffold. In comparison with scattered microvessel-like structures on random scaffolds, aligned microvessel-like structures were observed on aligned scaffolds. We firstly indicated that aligned electrospun fibers is sufficient to align microvessel-like structures formed by co-culturing the HUVECs and iPSC-MSCs, which provides a basis for further constructing diverse tissues containing aligned microvessel-like structures in vitro.