Thrombin induces endothelin expression in arterial smooth muscle cells

Thrombin has been shown to stimulate endothelin release by endothelial cell s, but the ability of thrombin to induce endothelin in nonendothelial cells is less well-known. Incubation of rat aortic smooth muscle cells with thro mbin resulted in a stimulation of preproendothelin-1 (preproET-1) mRNA expr ession. This induction of preproET-1 mRNA expression by thrombin was accomp anied by the release of immunoreactive peptide ET-1 into the extracellular medium. The synthetic thrombin receptor activator peptide (TRAP) confirmed ligand-specific receptor action to induce preproET-1 mRNA. Nuclear run-on a nalysis revealed that the transcriptional rate of preproET-1 mRNA increases twofold after 1 h of incubation with thrombin. In cells treated with throm bin, the half-life of preproET-1 mRNA was identical to that in untreated co ntrol cells. These results demonstrated that thrombin regulates endothelin synthesis at a transcriptional level but does not influence mRNA stability. Inhibition of protein kinase C (PKC) with selective inhibitors (chelerythr ine and bisindolylmaleimide I) before thrombin stimulation failed to signif icantly inhibit preproET-1 gene expression. Inhibition of mitogen-activated protein (MAP) kinase kinase and protein tyrosine kinase decreased preproET -1 mRNA expression in thrombin-stimulated smooth muscle cells. Furthermore, addition of an activator of peroxisome proliferator-activated receptors-al pha (PPAR alpha), fenofibrate, prevented the preproET-1 gene induction in r esponse to thrombin. These results demonstrated that thrombin-induced endot helin gene transcription involved MAP kinase kinase rather than the PKC cas cade in smooth muscle cells, which was repressed by PPARa stimulation.