Fluid flow decreases mI qA levels and peptide release in endothelial cells preproendothelin suppresses endothelin-1 cultured human

Eedothelin-1, a 21-amino acid peptide secreted by endothelial cells, has constrictor and mitogenic activity for vascular smooth muscle cells, and its mitogenic activity is synergistic with that of platelet-derived growth factor. Endothelial cell-derived endothelin-1 might therefore contribute to intimal hyperplasia in reendothelialized segments of vascular grafts or of endarterectomy and angioplasty sites. Because intimal hyperplasia occurs most often at sites with disordered flow patterns and lower fluid shear stress, we tested the effects of static culture versus high laminar shear stress (25 dyne/cm 2) on endothelin-1 precursor (preproendothelin) gene mRNA transcript levels and endothelin-1 peptide release in cultured human endothelial cells. Primary cultures of human umbilical vein endothelial cells were subje.cted to controlled levels of shear stress in parallel plate flow chambers for 24 hours. To dietect preproendothelin mRNA we applied a linked reverse transcriptasepolymerase chain reaction (RT/PCR) to RNA extracted from cultures. Southern blots of RT/PCR reaction products were hybridized with radioactive phosphorous (a2p) labeled probes for the amplified preproendothelin complementary deoxyribonucleic acid (cDNA). Detection by RT/PCR ofmRNA for glyceraldehyde 3-phosphate dehydrogenase was used to measure a c.onstitutively expressed control signal. Endothelin-1 release into cultttre medium was measured by radioimmunoassay. Application of 25 dyne/era 2 of shear stress for 24 hours sharply reduced endothelial cell levels of precursor preproendothelin mRNA. High shear stress also decreased endothelin-1 peptide release rates into culture medium from 0.55 0.02 and 0.49 + 0.03 ng/106 endothelial cells/hour (n = 3 replicate cultures in each experiment) (static conditions) to values of 0.13 +0.03 and 0.28 + 0.06 ng/106 cells/hour (shear stress) (p < 0.01). The flow-induced suppression of preproendothelin gene expression suggests that endothelin-1 release from endothelial cells might help mediate the inverse relationship between local fluid shear rates and the severity of neointimal hyperplasia. (J VAse SvRa 1991;14:1-9.)