In vitro investigations of the responses of vascular endothelium to fluid shear stress have typically been conducted under conditions where the time-mean shear stress is uniform. In contrast, the in vitro experiments reported here have re-created the large gradients in surface fluid shear stress found near arterial branches in vivo; specifically, we have produced a disturbed-flow region that includes both flow separation and reattachment. Near reattachment regions, shear stress is small but its gradient is large. Cells migrate away from this region, predominantly in the downstream direction. Those that remain divide at a rate that is high compared with that of cells subjected to uniform shear. We speculate that large shear stress gradients can induce morphological and functional changes in the endothelium in regions of disturbed flow in vivo and thus may contribute to the formation of atherosclerotic lesions.