Abstract 1332: Fluid Shear Stress-Dependent Activation of the AMP-Activated Protein Kinase Attenuates Ras-Dependent Signaling by Inhibiting the Activity and Expression of the Hydroxy-Methylglutaryl Coenzyme A Reductase in Endothelial Cells
The hydroxy-methylglutaryl coenzyme A reductase (HCR) is the rate-limiting enzyme for cholesterol synthesis and is phosphorylated and inactivated by the AMP-activated kinase (AMPK). As shear stress activates the AMPK in endothelial cells we determined whether it affects HCR activity and subsequent HCR-dependent signaling. HCR activity in microsomes isolated from cultured endothelial cells exposed to shear stress (12 dynes cm-2, 2 to 24 hours) or the AMPK activator AICAR, was significantly decreased compared to controls. Inhibition of the endothelial nitric oxide (NO) synthase (eNOS) with Nω-nitro L-arginine (300 μM) attenuated the shear stress-dependent activation of AMPK as well as the decrease in HCR activity. Exposure of endothelial cells to fluid shear stress also decreased the expression of HCR mRNA, an effect that was attenuated after inhibition of the eNOS as well as after inhibition of the guanylyl cyclase with NS2028, again indicating an NO-mediated mechanism. In femoral arteries from wild-type mice provided with a running wheel for voluntary exercise (3838±348 m/day for 7 days) HCR mRNA expression was decreased compared to sedentary mice. Moreover, in liver microsomes from eNOS knock-out mice HCR activity was significantly higher than in microsomes from wild-type mice, suggesting that NO plays a role in the regulation of HCR activity. As the pleiotropic effects of statins have been attributed to the inhibition of protein isoprenylation we compared the effects of shear stress and cerivastatin on endothelial cell activation by bradykinin. Fluid shear stress (18 hours) and cerivastatin (3 nM, 18 hours) both attenuated the bradykinin-induced activation of Ras as well as the phosphorylation and activation of ERK1/2. These data indicate that fluid shear stress regulates endothelial cell HCR activity and Ras-dependent signaling via short- and long-term mechanisms involving NO and the AMPK.