Molecular mechanisms of endothelin-1-induced cell-cycle progression - Involvement of extracellular signal-regulated kinase, protein kinase C and phosphatidylinositol 3-kinase at distinct points

Although it is well established that endothelin-1 (ET-1) has not only vasoconstrictive effects but also mitogenic effects, which seem to be implicated in vascular remodeling, little is known about the molecular mechanisms by which ET-I induces cell-cycle progression. In this study, we examined the effects of ET-1 on the cell-cycle regulatory machinery, including cyclins, cyclin-dependent kinase (cdk), and cdk inhibitors in NIH3T3 cells. ET-1 increased cyclin D1 protein (5.1 +/- 1.9-fold increase, 8 hours after stimulation, P<0.05), cdk4 kinase activity (2.8 +/- 0.5-fold increase, 12 hours after stimulation, P<0.01), and cdk2 kinase activity (2. +/- 0.4-fold increase, 16 hours after stimulation, P<0.05) in a time- and dose-dependent manner. ET-l-induced increase in cyclin D1 protein, and cdk4 kinase activity was not significantly inhibited by an inhibitor of the mitogen-activated protein kinase kinase 1/2, PD98059, nor by the protein kinase C inhibitor calphostin C, whereas ET-l-induced upregulation of cyclin D1 protein and cdk4 kinase activity was significantly inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002. In contrast, ET-1-induced activation of cdk2 kinase was significantly inhibited by PD98059, calphostin C, and LY294002. ET-1 increased H-3-thymidine uptake in a time-dependent fashion (0 hours, 4216 +/- 264 cpm per well; 8 hours, 5025 +/- 197 cpm per well; 16 hours, 9239+/-79 cpm per well, P<0.001 versus 0 hours). ET-1-induced increase in 3H-thymidine uptake was significantly inhibited by PD98059, calphostin C, and LY294002. These results suggest that ET-l-induced cell-cycle progression is, at least in part, mediated by the extracellular signal-regulated kinase, protein kinase C, and phosphatidylinositol 3-kinase and that those pathways may be involved in the progression of the cell cycle at distinct points.