Cathepsins, the lysosomal cysteine proteases are involved in vascular remodeling and atherosclerosis. Genetic knockout of cathepsins S and K in mice has shown to reduce atherosclerosis, although the molecular mechanisms remain unclear. Since atherosclerosis preferentially occurs in arteries exposed to disturbed flow conditions, we hypothesized that shear stress would regulate cathepsin K expression and activity in endothelial cells. Mouse aortic endothelial cells (MAEC) exposed to proatherogenic oscillatory shear (OS, ± 5 dyn/cm² for 1 day) showed significantly higher cathepsin K expression and activity than that of atheroprotective, unidirectional laminar shear (LS, 15 dyn/cm² for 1 day). Western blot and active-site labeling studies showed an active, mature-form of cathepsin K in the conditioned medium of MAEC exposed to OS, but not in that of LS. Functionally, MAEC exposed to OS significantly increased elastase and gelatinase activity above that of LS. The OS-dependent elastase and gelatinase activities were significantly reduced by knocking-down cathepsin K with siRNA, but not by a non-silencing siRNA control, suggesting that cathepsin K is a shear-sensitive protease. In addition, immunohistochemical analysis of atherosclerotic human coronary arteries showed a positive correlation between the cathepsin K expression levels in endothelium and elastic lamina integrity. These findings suggest that cathepsin K is a mechanosensitive, extracellular matrix protease, which, in turn, may be involved in arterial wall remodeling and atherosclerosis.