Fluid dynamics strongly influences endothelial cell function, and participates in the localization of atherosclerotic plaques at blood vessel branches. We investigated the hypothesis that wild-type human aortic endothelial cells (HAEC) exposed to prolonged pulsatile flow stimulation have levels of phosphorylated mitogen-activated protein kinases (MAPK) that are significantly greater than those observed in statically grown cultures. HAEC were exposed to pulsatile laminar shear stress in a parallel-plate flow chamber and analyzed for levels of phosphorylated ERK, JNK and p38 at 1, 10 and 20 h. While some MAPK exhibited alternating patterns of phosphorylation, others were characterized by steady increases or unchanged profiles until the terminal (20 h) time point. However, at 20 h, each MAPK demonstrated an increase in phosphorylation versus statically cultivated cells. Further, 20 h cultures from 10 dyn/cm(2) pulsatile shear stress had higher levels of phosphorylation for each MAPK than those from 2 dyn/cm(2). The finding that MAPK species can be phosphorylated in response to a prolonged pulsatile shear stress in both a time and magnitude dependent manner is an interesting result that may help to explain how the differential behaviors observed between cells from different flow environments can be generated and maintained.