Endothelial cells (ECs) and smooth muscle cells (SMCs) are the most crucial components of the vascular wall, and the interactions between them are essential for the maintenance of normal vascular physiology, as well as the initiation and development of cardiovascular diseases. Various typical co-culture models have been developed to simulate the location and growth situation of ECs and SMCs in vivo. In this review, the co-culture systems combined with the device applying fluid shear stress were introduced with discussion on the strengths and limitations of each system. The influences of ECs-SMCs interaction on the phenotype and alignment of ECs and SMCs, growth and migration of SMCs, and adhesion molecules expression of ECs under shear stress were briefly reviewed. The established evidence indicates that nitric oxide (NO), cytokines and microRNA are the most important signal molecules mediating the interactions between ECs and SMCs.