Apoptotic hepatocellular carcinoma HepG2 cells accelerate blood coagulation

Backgrounds and Aim: Intrasinusoidal microthrombosis is considered to be a cause of massive hepatocyte death in fulminant hepatic failure. Generally, apoptotic cells express phosphatidyl serine (PS) outside the plasma membrane, which is also expressed on the surface of activated platelets and accelerates fibrin-thrombus formation. Therefore, the acceleration of blood coagulation on the surface of apoptotic hepatocytes may occur because hepatocytes are in direct contact with plasma that passes through fenestrations of the sinusoidal endothelium. To test this hypothesis, we investigated the coagulation activity of apoptotic hepatocytes. Methods: (1) The apoptosis of Hep G2 cells was induced by staurosporin (STS). PS expression was determined by confocal microscopy with FITC-annexin V, a specific inhibitor of PS activity. (2) One million HepG2 cells treated with STS with or without pretreatment with annexin V were exposed to activated factor X, CaCl(2) and prothrombin. Thrombin generation was determined using a thrombin-specific chromogenic substrate. Results: (1) The percentage of apoptotic cells and PS-expressing cells markedly increased following the STS-treatment. (2) The thrombin generation significantly increased with STS-treatment in a dose dependent manner. (3) The increase in thrombin generation by STS-treatment was abolished following pretreatment with annexin V. Conclusion: Apoptotic hepatocytes accelerate blood coagulation through the expression of a PS-dependent pro-coagulant surface.