FLOW-MEDIATED REGULATION OF ENDOTHELIN RECEPTORS IN COCULTURED VASCULAR SMOOTH-MUSCLE CELLS - AN ENDOTHELIUM-DEPENDENT EFFECT

The objective of this study was to determine the effect of pulsatile f low on endothelin (ET) receptor expression in vascular smooth muscle c ells (VSMC) cocultured with endothelial cells (EC). Using a perfused t ranscapillary coculture system which permits the chronic exposure of c ultured EC and VSMC to physiological shear stresses, cocultures were e xposed to stepwise increases in flow up to (1) 2 ml/min (low flow: 0.5 dyn/cm(2)) or (2) 44 ml/min (high flow: 15 dyn/cm(2)) and maintained for 72 h before SMC and EC were harvested separately. There was a sign ificant increase in [I-125]-ET-1 binding in cocultured VSMC exposed to high flow as compared to low flow (B-max,,: 75 +/- 22 vs. 152 +/- 10 fmol [I-125]ET-1 bound/mg protein) in the absence of any change in the affinity (K-D) of ET-1 for its receptor. ET-I peptide mRNA levels wer e significantly decreased in EC exposed to high flow. The increase in [I-125]ET-I binding was associated with an increase in ET-A and ET-B r eceptor mRNA levels and was EC dependent as [I-125]ET-1 binding in mon ocultured VSMC was the same, regardless of flow conditions. However, t he amount of [I-125]ET-1 binding on VSMC cultured in the absence of EC was significantly greater than that on cocultured VSMC. High flow cau sed a significant increase in endothelial nitric oxide synthase (NOS) activity in EC and prostacyclin levels in the perfusing medium. Flow-m ediated upregulation of ET receptors was diminished by treatment with N-G-nitro-L-arginine-methyl ester, a NOS inhibitor, whereas indomethac in, a cyclooxygenase inhibitor, had no significant effect. Collectivel y, these data suggest that flow-induced changes in ET receptor express ion in VSMC are endothelium dependent and are in part mediated by nitr ic oxide. Modulation of ET receptor expression by EC may thus represen t an important mechanism whereby hemodynamic forces regulate vessel wa ll function.