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Shear Stress Causes Nuclear Localization of Endothelial Glucocorticoid Receptor and Expression From the GRE Promoter

From the Institute for Medicine and Engineering, Departments of Bioengineering (J.Y.J., H.J., S.L.D.) and Chemical and Biomolecular Engineering (S.L.D.), University of Pennsylvania, Philadelphia, Pa.

Correspondence to Dr Scott L. Diamond, Institute for Medicine and Engineering, 1010 Vagelos Research Laboratory, 3340 Smith Walk, Philadelphia, PA 19104. E-mail sld{at}seas.upenn.edu

We tested the hypothesis that steady laminar shear stress activates the glucocorticoid receptor (GR) and its transcriptional signaling pathway in an effort to investigate the potential involvement of GR in shear stress-induced antiatherosclerosis actions in the vasculature. In both bovine aortic endothelial cells (BAECs) and NIH3T3 cells expressing GFP-GR chimeric protein, wall shear stress of 10 or 25 dynes/cm² caused a marked nuclear localization of GFP-GR within 1 hour to an extent comparable to induction with 25 µmol/L dexamethasone. The shear mediated nuclear localization of GFP-GR was significantly reduced by 25 µmol/L of the MEK1 inhibitor (PD098059) or the PI 3-kinase inhibitor (LY294002). Also, Western blots demonstrated translocation of endogenous GR into nucleus of sheared BAECs. Promoter construct studies using glucocorticoid response element (GRE)-driven expression of secreted alkaline phosphatase (SEAP) indicated that BAECs exposed to shear stress of 10 and 25 dynes/cm² for 8 hours produced >9-fold more SEAP (n=6; P<0.005) than control cells, a level comparable to that observed with dexamethasone. Shear stress enhanced SEAP expression at 6 hours was reduced 50% (n=5; P<0.005) by MEK1/2 or PI 3-kinase inhibitors, but not by the NO inhibitor, L-NAME. Finally, in human internal mammary artery, endothelial GR is found to be highly nuclear localized. We report a new shear responsive transcriptional element, GRE. The finding that hemodynamic forces can be as potent as high dose glucocorticoid steroid in activating GR and GRE-regulated expression correlates with the atheroprotective responses of endothelial cells to unidirectional arterial shear stress.

Key Words: endothelium • shear stress • hemodynamics • glucocorticoid receptor

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