Nitric oxide-dependent and nitric oxide-independent transcriptional responses to high shear stress in endothelial cells.
Shear stress modulates gene expression in endothelial cells (ECs) partly through nitric oxide (NO), acting via enhanced cGMP formation by guanylyl cyclase (GC). We addressed non-cGMP-mediated transcriptional responses to shear stress in human umbilical ECs subjected to high-laminar shear stress (25 dyn/cm2; 150 minutes). RNA was isolated, reverse-transcribed, Cy3/5-labeled, and hybridized to 19 K human microarrays. High shear (n=6), high shear with 100 micromol/L L-NAME (n=3), and high shear with 10 micromol/L ODQ (GC inhibitor) in the perfusate (n=3) was compared with samples not subjected to flow. Among genes responding to high shear were HMOX1 (up) and PPARG (down). A high percentage of gene expression modulation by shear was absent during concomitant L-NAME or ODQ. Several transcriptional modulators were found (up: SOX5, SOX25, ZNF151, HOXD10; down: SOX11); a number of genes were regulated by shear and by shear with ODQ, but not regulated during L-NAME, indicating a nitric oxide synthase (NOS)-dependent, guanylyl cyclase (GC)-independent pathway. Several genes only responded to shear stress during L-NAME, others only responded to shear during ODQ. Upstream binding site analysis indicated shear stress and NO-dependent regulation of transcription via SOX5 and SOX9. Although NO importantly modulated the effect of shear stress on EC transcription, HMOX1 was consistently induced by shear stress, but not dependent on NOS or GC. Using bio-informatics software and databases, a promoter analysis identified SOX5 and SOX9 as potential, novel, shear-sensitive, and NO-dependent transcriptional regulators. The role of HMOX1 as a potential backup for NOS and the downstream role of SOXes should be explored.
Braam
Branko
B
Department of Nephrology and Hypertension, University Medical Center, F03.226, P.O. Box 85500, 3508 GA Utrecht, The Netherlands. g.b.braam@azu.nl
de Roos
Remmert
R
Bluyssen
Hans
H
Kemmeren
Patrick
P
Holstege
Frank
F
Joles
Jaap A
JA
Koomans
Hein
H
D001665:Binding Sites; D006152:Cyclic GMP / Q000502:physiology; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000502:physiology; D051766:Early Growth Response Protein 1; D042783:Endothelial Cells / Q000378:metabolism / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors; D006419:Heme Oxygenase (Decyclizing) / Q000096:biosynthesis; D051547:Heme Oxygenase-1; D006609:High Mobility Group Proteins / Q000502:physiology; D006801:Humans; D017874:Immediate-Early Proteins / Q000096:biosynthesis; D008565:Membrane Proteins; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D052250:Nitric Oxide Synthase Type III; D009687:Nuclear Proteins / Q000502:physiology; D010069:Oxadiazoles / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D055758:SOX9 Transcription Factor; D055755:SOXD Transcription Factors; D013314:Stress, Mechanical; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology; D014158:Transcription, Genetic / Q000502:physiology*
C095284:1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D004791:Enzyme Inhibitors; D006609:High Mobility Group Proteins; D017874:Immediate-Early Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; D010069:Oxadiazoles; D011810:Quinoxalines; C490301:SOX5 protein, human; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; D055755:SOXD Transcription Factors; D014157:Transcription Factors; D009569:Nitric Oxide; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C496541:HMOX1 protein, human; D006419:Heme Oxygenase (Decyclizing); D051547:Heme Oxygenase-1; D006162:Guanylate Cyclase; D006152:Cyclic GMP; D019331:NG-Nitroarginine Methyl Ester