Effect of a phorbol ester on immunoreactive endothelin-1 release from cultured porcine aortic endothelial cells

This study was designed to examine how protein kinase C (PKC) regulates the release of endothelin-1 (ET-1) from cultured porcine aortic endothelial cells. We measured the release of immunoreactive (IR)-ET-1 from cells cultured for up to 72 h in the presence or absence of a phorbol ester TPA. The release of IR-ET-1 from control cells (no TPA) increased according to time for up to 72 h. In the presence of TPA, the release of IR-ET-1 from the cells was higher than the control level for up to 8 h, but was lower thereafter and reached a plateau after 48 h. TPA dose-dependently stimulated IR-ET-1 release during incubation for 4 h, but suppressed it after incubation for 72 h. Stimulation of PKC by diacylglycerol mimicked the early (4 h) action of TPA. On the other hand, pretreatment of cells with TPA to downregulate PKC significantly suppressed basal and thrombin- or FCS-stimulated IR-ET-1 release. These findings suggest that the activation of PKC is related to the stimulation of ET-1 release and that down-regulation of PKC leads to the suppression of ET-1 release from cultured endothelial cells.