Combined effect of retinoic acid and basic FGF on PAI-1 gene expression in vascular smooth muscle cells

Objective: Aberrant regulation of the synthesis and degradation of the extracellular matrix (ECM) is associated with the pathophysiology of vascular disease. Plasminogen activator inhibitor-1 (PAI-1) plays a crucial role in regulating the quantity and composition of ECM. However. regulatory mechanisms underlying the expression of the PAI-1 gene remain unclear. We examined the effects of all-trans-retinoic acid (atRA), either alone or in combination with mitogenic growth factor, basic fibroblast growth factor (bFGF), on the PAI-1 expression in cultured vascular smooth muscle cells (SMCs). Methods: Cultures of the rabbit vascular smooth muscle cell line C2/2 were used to study the effects: of atRA and bFGF separately or together. Results: Treatment of vascular SMCs with atRA in combination with bFGF resulted in an additional increase in PAI-1 expression both at the mRNA and protein levels. In contrast. tissue-type plasminogen activator. urokinase-type plasminogen activator and tissue factor mRNA levels were only minimally affected. The all-trans-RA- and bFGF-mediated increases in PAI-1 mRNA levels were markedly attenuated by the tyrosine kinase inhibitor genistein. but not by MEK1 or p38MAP kinase inhibitors. The rate of decrease in PAI-1 mRNA levels after actinomycin D treatment was not affected by atRA and bFGF. Transient transfection of the PAI-1 promoter-luciferase reporter gene, which contains 967 bp of the 5'-flanking region of the human PAI-1 gent, revealed that atRA and bFGF additionally increased transcription from this promoter. progressive 5'-deletion revealed that the promoter region required for such an effect lies between -967 and -260, which contains no canonical sequence for the RA-response element. In agreement with the role of PAI-1 in the inhibition of fibrinolytic activity which stimulates ECM degradation, cell migration was inhibited by treatment with atRA and bFGF. Conclusions: These results indicate that atRA and bFGF can function in a combined fashion and induct PAI-1 synthesis in vascular SMCs, and suggest a role for these two compounds in regulating SMC migration. (C) 2001 Elsevier Science B.V. All rights reserved.