Heparanase induces tissue factor pathway inhibitor expression and extracellular accumulation in endothelial and tumor cells

Abstract

SummaryHeparanase activity is implicated in cell invasion, tumor metastasis and angiogenesis. Recently, we have reported that heparanase stimulates tissue factor (TF) expression in endothelial and cancer cells, resulting in elevation of coagulation activity. We hypothesized that heparanase regulates other coagulation modulators, and examined the expression and localization of tissue factor pathway inhibitor (TFPI) following heparanase over-expression or exogenous addition. Primary human umbilical vein endothelial cells (HUVEC) and human tumor-derived cell lines were incubated with heparanase, or were stably transfected with heparanase gene-constructs, and TFPI expression and secretion were examined. Heparanase over-expression or exogenous addition stimulated TFPI expression by 2–3 folds. TFPI accumulation in the cell culture medium exceeded in magnitude the observed induction ofTFPI gene transcription reaching 5– to 6-fold increase. Extracellular accumulation of TFPI was evident already 60 min following heparanase addition, prior toTFPI protein induction, and correlated with increased coagulation activity. This effect was found to be independent of heparanase enzymatic activity and interaction with heparan-sulfate, and correlated with reduced TFPI levels on the cell surface. Data were verified in heparanase transgenic mice tissues and plasma. Interaction between heparanase and TFPI was evident by co-immunoprecipitation. Interaction of heparanase with TFPI resulted in its displacement from the surface of the vascular endothelium and in increased pro-coagulant activity. Thus, heparanase facilitates blood coagulation on the cell surface by two independent mechanisms:dissociation ofTFPI from the vascular surface short after local elevation of heparanase levels, and subsequent induction of TF expression.

Heparanase Regulates Thrombosis in Vascular Injury and Stent-Induced Flow Disturbance

“…In parallel, heparanase also appears to stimulate TF pathway inhibitor (21). Additionally, exogenously applied heparanase has been shown to modify in vitro coagulation on endothelial cells in culture (20,22).…”

Insights into the key roles of proteoglycans in breast cancer biology and translational medicine

“…Heparanase function however is not limited solely to its enzymatic activity. Enzymatically inactive heparanase can activate signaling molecules such as AKT and p38 [256, 257] and promote transcription of several biologically important effectors [ e.g., hepatocyte growth factor (HGF) and tissue factor] [258, 259]. This implies heparanase has broad functions beyond its impact on HS.…”

Heparanase and Coagulation–New Insights

“…There are also reports on increased plasma levels of TFPI in relation to diabetes mellitus,23 renal diseases,24 and cancer 25,26. Recently we demonstrated that exogenous addition or over-expression of heparanase by transfected cells resulted in release of TFPI from the cell surface and its accumulation in the cell culture medium 27. Importantly, the in vitro studies were supported by elevation of TFPI levels in the plasma of transgenic mice over-expressing heparanase.…”

“…Heparanase was recently revealed as an important modulator of blood coagulation 13,17,18,27. Generating peptides inhibiting heparanase procoagulant activity36 strengthens the role of heparanase as a new procoagulant factor in the coagulation cascade.…”

“…Both functions require secretion of heparanase, but not its enzymatic activity. The underlying mechanism is apparently release of TFPI due to its physical interaction with the secreted heparanase, as clearly evident by co-immunoprecipitation experiments,27 reflecting a functional interaction between heparanase and a membrane protein.…”