Shear stress causes nuclear localization of endothelial glucocorticoid receptor and expression from the GRE promoter.
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 micromol/L dexamethasone. The shear mediated nuclear localization of GFP-GR was significantly reduced by 25 micromol/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.
Ji
Julie Y
JY
Institute for Medicine and Engineering, Department of Bioengineering, University of Pennsylvania, Philadelphia, Pa 19104, USA.
Jing
Huiyan
H
Diamond
Scott L
SL
D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D002417:Cattle; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D008323:Mammary Arteries / Q000166:cytology / Q000378:metabolism; D051379:Mice; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology; D013314:Stress, Mechanical; D015533:Transcriptional Activation / Q000502:physiology
D004791:Enzyme Inhibitors; D008164:Luminescent Proteins; D011965:Receptors, Glucocorticoid; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D003907:Dexamethasone; D019331:NG-Nitroarginine Methyl Ester