Genistein reduces tumor necrosis factor alpha-induced plasminogen activator inhibitor-1 transcription but not urokinase expression in human endothelial cells.
The plasminogen activator inhibitor PAI-1 is markedly elevated in vivo and in vitro upon exposure to the inflammatory mediators tumor necrosis factor alpha (TNF alpha), interleukin-1 (IL-1), and bacterial lipopolysaccharide. Here we report that the isoflavone compound genistein prevents the increase in synthesis of PAI-1 induced by these inflammatory mediators in human endothelial cells in vitro, and partially reduces the basal PAI-1 production by these cells. These effects of genistein were accompanied by a decrease in PAI-1 mRNA and in a suppression of the PAI-1 transcription rate as shown by run-on assay. A specific action of genistein, probably by inhibiting a tyrosine protein kinase, is likely, because the structural genistein analogue daidzein, which has a low tyrosine protein kinase inhibitor activity, did not inhibit PAI-1 synthesis. Vanadate, a tyrosine protein phosphatase inhibitor, increased PAI-1 production. The effect of genistein on PAI-1 synthesis was rather selective. Herbimycin A also reduced PAI-1 synthesis, but several other tyrosine protein kinase inhibitors, namely tyrphostin A47, methyl-2,5-dihydroxy-cinnamate, and compound 5, were unable to do so. All these tyrosine protein kinase inhibitors reduced basic fibroblast growth factor (b-FGF)-induced [3H]thymidine incorporation in endothelial cells. This indicates that the effect of genistein on PAI-1 transcription proceeds independently of its effect on mitogenesis. In contrast to TNF-alpha-induced PAI-1 production, the transcription and synthesis of urokinase-type plasminogen activator (u-PA) was not inhibited by genistein. A TNF-alpha-mutant (Trp32Thr86TNF alpha) that specifically recognizes the 55-kD TNF-receptor, mimicked the effects of TNF alpha on both PAI-1 and u-PA. Because genistein affected PAI-1, but not u-PA induced by this mutant, involvement of different TNF-receptors cannot underlie the difference in the effects of genistein on PAI-1 and u-PA synthesis. Because genistein also inhibited PAI-1 induction by thrombin and IL-4, it is likely that genistein does not act on a TNF alpha-receptor-coupled protein kinase but on the signal transduction pathway enhancing PAI-1 transcription. Our results suggest that the TNF alpha-induced signal transduction pathway of PAI-1 transcription involves a genistein-sensitive step that is not involved in the induction of u-PA by TNF alpha. Given the limited sensitivity to several other tyrosine protein kinase inhibitors, this genistein-sensitive step may be a potential target for pharmacologic intervention to reduce elevated plasma PAI-1 levels.
van Hinsbergh
V W
VW
Gaubius Laboratory TNO-PG, Leiden, The Netherlands.
Vermeer
M
M
Koolwijk
P
P
Grimbergen
J
J
Kooistra
T
T
D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D019833:Genistein; D006801:Humans; D066298:In Vitro Techniques; D007529:Isoflavones / Q000494:pharmacology*; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D018124:Receptors, Tumor Necrosis Factor / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D007529:Isoflavones; D017395:Plasminogen Activator Inhibitor 1; D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D016222:Fibroblast Growth Factor 2; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator