Quantitation and properties of the active and latent plasminogen activator inhibitors in cultures of human endothelial cells.
Human endothelial cells release two forms of a plasminogen activator-specific inhibitor: an active form that readily binds to and inhibits plasminogen activators and an inactive or latent form that has no anti-activator activity but which can be activated by denaturation. Latent and active forms of plasminogen activator-specific inhibitor were measured in cultures of human umbilical vein endothelial cells. Latent inhibitor was activated by treatment with 1% sodium dodecyl sulfate (SDS), and both forms were assayed by the 125I-fibrin plate method. After 16 hours, the conditioned medium contained 104.6 U/mL latent inhibitor activity and 2.6 U/mL active inhibitor. The level of each form in the culture medium increased with time although the activity associated with the latent form rose more rapidly: the ratio of latent to active inhibitor activity was 12 at four hours (10.3 U/mL v 0.86 U/mL) and reached 56 at 24 hours (155.3 U/mL v 2.80 U/mL). Intracellular inhibitor activity was associated with the active form only; no additional inhibitor activity was observed following SDS treatment of cell extracts. A decline in active inhibitor activity occurred during incubation at 37 degrees C with a 50% reduction in activity occurring in two hours. Treatment of conditioned medium with 10 U/mL thrombin also resulted in a loss of active inhibitor activity. The latent inhibitor, however, was not affected by either of these conditions. The inhibitor activity lost during incubation at 37 degrees C or thrombin treatment could be regenerated by SDS treatment, suggesting that the loss of the active inhibitor activity represented a conversion of this form to its latent counterpart. Thus, the concentration, stability, and regulation of these two forms of plasminogen activator inhibitor in human endothelial cell cultures differ significantly.
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D002478:Cells, Cultured; D004727:Endothelium / Q000032:analysis*; D005260:Female; D005337:Fibrin; D006801:Humans; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D011247:Pregnancy; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013997:Time Factors; D010959:Tissue Plasminogen Activator / Q000037:antagonists & inhibitors*; D014471:Umbilical Veins / Q000166:cytology; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors*
D012967:Sodium Dodecyl Sulfate; D011092:Polyethylene Glycols; D005337:Fibrin; D017830:Octoxynol; D013917:Thrombin; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator