An endothelial cell (EC) monolayer aligned along the direction of blood flow in vivo shows excellent capacity for anti-inflammation and anti-thrombosis. Therefore, aligned electrospun fibers have been much studied in the field of vascular implants since they are considered to facilitate the formation of an aligned EC monolayer, yet few research studies have been comprehensively reported concerning the effects of diameter scales of aligned fibers. In the present work, a series of aligned polycaprolactone (PCL) electrospun fibers with varying diameters ranging from dozens of nanometers to several micrometers were developed, and the effects of the fiber scales on EC behaviors, hemocompatibility as well as inflammatory cell behaviors were investigated, to evaluate their potential performance in the field of vascular implants. Our results showed that platelets exhibited small attachment forces on all fibers, and the anticoagulation property improved with the decrease of the fiber diameters. The impact of fiber diameters on human umbilical vein endothelial cell (HUVEC) adhesion and NO release was limited, while significant on HUVEC proliferation. With the increase of the fiber diameters, the elongation of HUVECs on our samples increased first then decreased, and exhibited maximum elongation degrees on 2738 nm and 2036 nm due to the strong contact guidance effect on these graphical cues; too thick or too fine fibers would weaken the contact guidance effect. Furthermore, we hypothesized that HUVECs cultured on 2036 nm had the smallest spreading area because of their elongation, but 2738 nm restricted HUVECs spreading limitedly. Similarly, NO production of HUVECs showed a similar change trend as their elongation degrees on different fibers. Except for 2036 nm, it exhibited the second highest NO production. For RAW 264.7 cells, poorer cell adhesion and lower TNF-α concentration of 1456 nm indicated its superior anti-inflammation property, while 73 nm showed a contrasting performance. Overall, these findings partly revealed the relationship between different topographies and cell behaviors, providing basic insight into vascular implant design.