title: Combined effect of retinoic acid and basic FGF on PAI-1 gene expression in vascular smooth muscle cells.
abstract: 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 gene, 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 induce PAI-1 synthesis in vascular SMCs, and suggest a role for these two compounds in regulating SMC migration.
authors.affiliation: Second Department of Internal Medicine, Gunma University School of Medicine, 3-39-15 Showa-machi, Maebashi, Gunma 371-8511, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Watanabe
authors.forename: M
authors.initials: M
authors.lastname: Kurabayashi
authors.forename: M
authors.initials: M
authors.lastname: Arai
authors.forename: K
authors.initials: K
authors.lastname: Sekiguchi
authors.forename: R
authors.initials: R
authors.lastname: Nagai
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D003720:Densitometry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D019833:Genistein / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D017395:Plasminogen Activator Inhibitor 1 / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000032:analysis*; D011817:Rabbits; D013925:Thromboplastin / Q000032:analysis / Q000235:genetics; D010959:Tissue Plasminogen Activator / Q000235:genetics; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000032:analysis / Q000235:genetics
chemical_list: D004791:Enzyme Inhibitors; D017395:Plasminogen Activator Inhibitor 1; D012333:RNA, Messenger; D016222:Fibroblast Growth Factor 2; D014212:Tretinoin; D013925:Thromboplastin; D019833:Genistein; D008156:Luciferases; D011505:Protein-Tyrosine Kinases; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator