Flow shear stress plays important roles in modulating differentiation of endothelial progenitor cells (EPCs). MicroRNAs are crucial for diverse cellular processes, but the expressions and functions of microRNAs in EPCs responding to mechanical stimuli remain unclear. We sought to determine the effects of microRNA-34a (miR-34a) and a novel target Forkhead box j2 (Foxj2) on shear stress-induced EPC differentiation. Human umbilical cord blood-derived EPCs were exposed to laminar shear stress of 15dyn/cm(2) with parallel plate flow chamber system. Real time RT-PCR showed that shear stress significantly increased miR-34a expression, which was accompanied by the endothelial differentiation of EPCs. Whereas Foxj2, a putative target of miR-34a predicted by multiple algorithms, was suppressed in this process. Dual luciferase reporter assays, as well as miR-34a mimics and inhibitor treatment were used to confirm the interplay between miR-34a and Foxj2. Our results revealed an inverse correlation of miR-34a and Foxj2 expressions implicated in the endothelial differentiation of EPCs. MiR-34a contributed to this process by up-regulating the expressions of endothelial cell markers, and down-regulating smooth muscular cell markers. In addition, Foxj2 overexpression attenuated endothelial differentiation of EPCs, while Foxj2 siRNA had the opposite effect. These data suggested a unique mechanism that shear stress induces the expression of miR-34a, which targets to Foxj2 and promotes endothelial differentiation of EPCs. The results provide new insights into miR-34a/Foxj2 on shear stress-induced EPC differentiation.