Tissue-type plasminogen activator (t-PA) is a potent and efficacious mitogen for growth-arrested cultured human aortic smooth muscle cells, stimulating an increase in cell number at 0.3-30 nM concentration. Double-chain t-PA is as efficient as single-chain t-PA in stimulating smooth muscle cell mitogenesis, whereas single-chain urokinase-type plasminogen activator (u-PA) or u-PA and plasmin or plasminogen are ineffective. Plasminogen activator inhibitor-1, Pefabloc-TPA, diisopropyl fluorophosphate or alpha 1-anti-trypsin inhibit the mitogenic effect of t-PA for smooth muscle cells in a dose-dependent manner, showing that it is dependent on the enzymatic activity. t-PA activated phosphoinositide turnover in smooth muscle cells through a pertussis toxin-insensitive pathway and stimulated proto-oncogene c-fos and c-jun mRNA levels. These findings indicate that t-PA stimulates vascular human smooth muscle cell proliferation and suggest for the first time that it may contribute to intimal smooth muscle cell proliferation after vascular injury as a result of angioplasty or vascular compromise during atherogenesis.