Down-regulation of neprilysin (EC3.4.24.11) expression in vascular endothelial cells by laminar shear stress involves NADPH oxidase-dependent ROS production.

Neprilysin (NEP, neutral endopeptidase, EC3.4.24.11), a zinc metallopeptidase expressed on the surface of endothelial cells, influences vascular homeostasis primarily through regulated inactivation of natriuretic peptides and bradykinin. Earlier in vivo studies reporting on the anti-atherosclerotic effects of NEP inhibition and on the atheroprotective effects of flow-associated laminar shear stress (LSS) have lead us to hypothesize that the latter hemodynamic stimulus may serve to down-regulate NEP levels within the vascular endothelium. To address this hypothesis, we have undertaken an investigation of the effects of LSS on NEP expression in vitro in bovine aortic endothelial cells (BAECs), coupled with an examination of the signalling mechanism putatively mediating these effects. BAECs were exposed to physiological levels of LSS (10 dynes/cm(2), 24h) and harvested for analysis of NEP expression using real-time PCR, Western blotting, and immunocytochemistry. Relative to unsheared controls, NEP mRNA and protein were substantially down-regulated by LSS (>or=50%), events which could be prevented by treatment of BAECs with either N-acetylcysteine, superoxide dismutase, or catalase, implicating reactive oxygen species (ROS) involvement. Employing pharmacological and molecular inhibition strategies, the signal transduction pathway mediating shear-dependent NEP suppression was also examined, and roles implicated for G beta gamma, Rac1, and NADPH oxidase activation in these events. Treatment of static BAECs with angiotensin-II, a potent stimulus for NADPH oxidase activation, mimicked the suppressive effects of shear on NEP expression, further supporting a role for NADPH oxidase-dependent ROS production. Interestingly, inhibition of receptor tyrosine kinase signalling had no effect. In conclusion, we confirm for the first time that NEP expression is down-regulated in vascular endothelial cells by physiological laminar shear, possibly via a mechanotransduction mechanism involving NADPH oxidase-induced ROS production.

Cyclic strain-mediated matrix metalloproteinase regulation within the vascular endothelium: a force to be reckoned with.

The vascular endothelium is a dynamic cellular interface between the vessel wall and the bloodstream, where it regulates the physiological effects of humoral and biomechanical stimuli on vessel tone and remodeling. With respect to the latter hemodynamic stimulus, the endothelium is chronically exposed to mechanical forces in the form of cyclic circumferential strain, resulting from the pulsatile nature of blood flow, and shear stress. Both forces can profoundly modulate endothelial cell (EC) metabolism and function and, under normal physiological conditions, impart an atheroprotective effect that disfavors pathological remodeling of the vessel wall. Moreover, disruption of normal hemodynamic loading can be either causative of or contributory to vascular diseases such as atherosclerosis. EC-matrix interactions are a critical determinant of how the vascular endothelium responds to these forces and unquestionably utilizes matrix metalloproteinases (MMPs), enzymes capable of degrading basement membrane and interstitial matrix molecules, to facilitate force-mediated changes in vascular cell fate. In view of the growing importance of blood flow patterns and mechanotransduction to vascular health and pathophysiology, and considering the potential value of MMPs as therapeutic targets, a timely review of our collective understanding of MMP mechanoregulation and its impact on the vascular endothelium is warranted. More specifically, this review primarily summarizes our current knowledge of how cyclic strain regulates MMP expression and activation within the vascular endothelium and subsequently endeavors to address the direct and indirect consequences of this on vascular EC fate. Possible relevance of these phenomena to vascular endothelial dysfunction and pathological remodeling are also addressed.