Antibodies to Thrombomodulin Induce Receptor-Mediated Endocytosis in Human Saphenous Vein Endothelial Cells

Abstract

SummaryThe membrane glycoprotein thrombomodulin (TM) is an essential endothelial cell (EC) cofactor, which forms a 1:1 stoichiometric complex with thrombin. Binding of thrombin to the high affinity TM receptor transforms its procoagulant activity into an anticoagulant potential, by activating protein C. The fate of TM in the presence of thrombin is still unclear: some authors claim that the thrombin-TM complex is internalized in EC, while others find this complex to be stable for at least 2 h at 37° C on the EC surface. In the present study, we investigated the interactions of thrombin and Fab-fragments of anti-TM antibodies, coupled to 5 or 15 nm gold particles with saphenous vein endothelial cells. Our results demonstrate that TM can be observed both on the plasma membrane and in coated structures only in the presence of anti-TM antibodies. Addition of thrombin decreased the extent of this labeling, while in double labeling experiments, where cells were incubated with 5 nm gold coupled thrombin and 15 nm gold coupled Fab fragments of anti-TM antibodies, thrombin was cointernalized only when anti-TM antibodies were present. These results show that thrombin-TM complex is not significantly internalized in EC. The internalization of this complex induced by anti-TM antibodies could play an important role in the thrombotic complications induced by anti-EC autoantibodies.

Increase in plasma thrombin-activatable fibrinolysis inhibitor may not contribute to thrombotic tendency in antiphospholipid syndrome because of inhibitory potential of antiphospholipid antibodies toward TAFI activation

“…IgG fractions of those serum samples inhibited the activation of protein C and bound to a recombinant version of the EGF-like portion of thrombomodulin. In addition, it has been demonstrated that the levels of expression of thrombomodulin on endothelial cells are reduced by internalization of the thrombomodulin-thrombin complex in the presence of the antibody to thrombomodulin [53].…”

Biomedical aspects of targeted delivery of drugs to pulmonary endothelium

“…Quite appropriate to its name, thrombomodulin acting in concert with proteins C and S converts thrombin into an antithrombotic enzyme that inactivates coagulation factors [67]. Hence, any intervention that may compromise thrombomodulin function, including targeting drugs that may block, inhibit or shed thrombomodulin, has the potential to shift the balance towards thrombosis [68,69]. Indeed, some of such interventions induced or aggravated thrombosis in animal models [48].…”

Purification and characterization of two forms of soluble thrombomodulin from human urine

“…The molecular and functional characteristics of membrane thrombomodulin have been well delineated [6, 71. Cell-surface expression of thrombomodulin is regulated by a number of mechanisms including a tumour-necrosisfactor-mediated process which regulates transcription of the thrombomodulin gene [ 81, internalization by receptor-mediated endocytosis [9] and proteolytic cleavage from the endothelial cell membrane [lo]. The relative role of each of these mechanisms in the surface expression of thrombomodulin has yet to be defined.…”