Shear stress magnitude and directionality modulate growth factor gene expression in preconditioned vascular endothelial cells.
OBJECTIVE
The purpose of this study was to simultaneously monitor the transcriptional levels of 12 endothelial growth factor genes in response to alterations in wall shear stress (WSS) under conditions relevant to the development of intimal hyperplasia, a major cause of arterial bypass graft failure.


METHODS
Human umbilical vein endothelial cells were preconditioned in vitro under steady flow (WSS, 15 dynes/cm(2)) for 24 hours before being subjected to WSS at 25 (Delta = +10), 15 (Delta = 0), 5 (Delta = -10), 2.5 (Delta = -12.5), and 0 (Delta = -15) dynes/cm(2) or low magnitude WSS reversal (-2.5 dynes/cm(2)) for 6 hours. A focused complementary DNA array was used to simultaneously measure messenger RNA expression levels for END1, endothelial nitric oxide synthase (NOS3), platelet-derived growth factor A, platelet-derived growth factor B (PDGFB), acidic fibroblast growth factor, basic fibroblast growth factor, transforming growth factor-alpha, transforming growth factor-beta, vascular endothelial growth factor, insulin-like growth factor-1, epidermal growth factor, and angiotensin converting enzyme.


RESULTS
Preconditioning significantly (P <.05) increased the fold expression of NOS3 (4.1 +/- 1.4), basic fibroblast growth factor (3.90 +/- 1.16), vascular endothelial growth factor (3.39 +/- 1.04), and insulin-like growth factor-1 (2.8 +/- 0.7) but decreased END1 (0.47 +/- 0.05) and PDGFB (0.70 +/- 0.04) messenger RNA expression levels relative to no-flow controls, an effect that was sustained on removal from flow for 6 hours. Notably, the ratio of END1/NOS3 expression was diminished (0.11 +/- 0.03) relative to that of cells maintained in static culture. Although few differences in gene expression from baseline (15 dynes/cm(2)) were measured in cells exposed to either constant (Delta = 0) or step decreases (Delta = -10, -12.5, or -15 dynes/cm(2)) in WSS, marked changes were seen in the group exposed to a step increase in WSS (Delta = +10) or to WSS reversal. Low magnitude retrograde WSS evoked significant (P <.05) transcriptional changes in multiple genes, including elevated END1 (4.1 +/- 0.5), platelet-derived growth factor A (1.5 +/- 0.2), PDGFB (2.3 +/- 0.3), and transforming growth factor-beta (1.5 +/- 0.2) levels, but depressed NOS3 (0.60 +/- 0.17) levels, and a marked increase in END1/NOS3 (6.7 +/- 1.6) when compared with equal magnitude antegrade WSS (2.5 dynes/cm(2)).


CONCLUSIONS
These results support the implementation of a preconditioning phase for in vitro WSS studies to establish a physiologic baseline. Our findings complement previous macroscale findings and are consistent with a cellular mechanism involving increased END1 and PDGFB levels, but decreased NOS3 levels, leading to intimal hyperplasia at regions of low magnitude reversing WSS.
Passerini
Anthony G
AG
Department of Biomedical Engineering, The University of Akron, OH, USA. passerin@mail.med.upenn.edu
Milsted
Amy
A
Rittgers
Stanley E
SE
D002478:Cells, Cultured; D016228:Endothelial Growth Factors / Q000032:analysis / Q000235:genetics*; D004730:Endothelium, Vascular / Q000166:cytology*; D004815:Epidermal Growth Factor / Q000032:analysis; D016222:Fibroblast Growth Factor 2 / Q000032:analysis; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000032:analysis; D036341:Intercellular Signaling Peptides and Proteins / Q000032:analysis; D008222:Lymphokines / Q000032:analysis; D019001:Nitric Oxide Synthase / Q000032:analysis; D052250:Nitric Oxide Synthase Type III; D007703:Peptidyl-Dipeptidase A / Q000032:analysis; D012313:RNA / Q000032:analysis; D033081:Shear Strength; D013314:Stress, Mechanical; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor; D012313:RNA; D007334:Insulin-Like Growth Factor I; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; D007703:Peptidyl-Dipeptidase A