Tumor necrosis factor induction of urokinase-type plasminogen activator in human endothelial cells

Abstract

Vascular endothelial cells undergo morphological and functional changes at sites of cell-mediated immune responses which may serve to promote the pathogenesis of inflammation. These changes, described as 'endothelial cell activation' can be invoked by a variety of cytokines which include interleukin I (IL-1), tumor necrosis factor (TNF), and lipopolysaccharide (LPS). We report here on the regulation of the plasminogen activator (PA) proteolytic system by human recombinant TNF alpha in short term cultures (less than 4 passages) of human umbilical vein endothelial cells (HUVECs). TNF alpha treatment of HUVECs enhanced the production of 55 kDa urokinase (u) PA activity and uPA antigen by fourfold, in a concentration dependent manner (5 - 100 U/ml), following a 24 h treatment as determined by PA zymography and micro-ELISA assays, respectively. This response was specific for uPA since, no change in extracellular tissue type PA activity and tPA antigen levels were noted under analogous conditions. A similar 4-fold increase in the de novo synthesis of [35S]-methionine radiolabeled uPA was observed by immunoprecipitation following a 24 h TNF treatment. The induction of uPA by TNF was inhibited by actinomycin D and cycloheximide implying the necessity of RNA and protein synthesis, respectively. The effect of TNF could not be prevented by the addition of IL-1 neutralizing antibodies. Therefore, it is unlikely that TNF acts through the induction of IL-I secretion. Time course studies using PA zymography indicate that within 8 h after TNF exposure, a 2-fold increase in uPA activity above untreated basal levels was observed. Upregulation of extracellular uPA production in HUVECs following TNF treatment suggests yet a new aspect of cellular and interstitital PA regulation in endothelium during inflammation and angiogenesis.