Activation of platelets in whole blood by recombinant factor VIIa by a thrombin‐dependent mechanism

Abstract

Summary. Using a diluted whole blood method of flow cytometric analysis, we have shown that platelets could be activated in vitro in the presence of high concentrations (100 nmol/l) of recombinant factor (F) VIIa (rFVIIa; NovoSeven Ò ) and 2AE5 mmol/l calcium chloride. This was demonstrated by a significant increase in the mean percentage of platelets expressing CD62P and their mean fluorescent intensity (MFI) after 30 min versus platelets incubated with calcium or rFVIIa alone or diluted blood alone. The presence of rFVIIa and calcium increased the exposure of the PAC-1 activation epitope of glycoprotein (Gp) IIb/IIIa. This effect was equally influenced by the presence of calcium alone but not by rFVIIa. The effect of rFVIIa was time and concentration dependent. Thrombin generation was also necessary, as the effect of rFVIIa was completely abrogated by the additional presence of hirudin. Furthermore, soy bean trypsin inhibitor (SBTI) but not corn trypsin inhibitor (CTI) abrogated CD62P exposure, suggesting that thrombin was derived via FX but not FXII activation. Exposure of CD62P demonstrated a significant lag phase, sometimes of the order of > 30 min, as well as large intersubject variation. Significant platelet activation was observed at a concentration as low as 25 nmol/l rFVIIa. Platelet-leucocyte aggregation was also increased in the presence of 25 nmol/l rFVIIa and calcium. No significant difference was observed between levels of CD62P in diluted whole blood and platelet-rich plasma adjusted to an identical platelet count after their exposure to rFVIIa and calcium for 30 min.

Solidification mechanisms of chitosan–glycerol phosphate/blood implant for articular cartilage repair

“…TF binding with trace amounts of activated FVIIa that are present in plasma 24 triggers the extrinsic clotting cascade and thrombin generation. In our in vivo experiments, we used a recombinant TFephospholipid preparation at concentrations previously employed in clinical applications for maxillary bone grafts 25 , rhFVIIa at concentrations around threefold greater than the target plasma levels for clinical applications (w2 mg/mL) 26 , and thrombin concentrations within the low range used in standard fibrin glue 27 . Our choice of these particular factors was thus based on their known ability to promote repair processes, as well as their current use in other clinical contexts which could help translate their use in cartilage repair strategies involving polymereblood implants.…”

Recombinant Factor VIIa Reduces Bleeding Risk in Patients on Platelet Aggregation Inhibitors Immediately prior to Renal Transplantation – A Retrospective Analysis

“…It must be taken into consideration that the in vivo bleeding time is not necessarily prolonged in drug-induced impairments of platelet functions. In our opinion, the effect of rFVIIa can be explained mainly by a disproportionately increased thrombin formation, which in turn leads to an enhanced inhibition of fibrinolysis via activation of thrombin activatable fibrinolysis inhibitor and also to an activation of platelets [18] . The absence of clinically relevant bleeding complications in our case series and the rapid postoperative onset of organ functioning suggests the conduct of studies with a bigger population.…”

Rapid reversal of coagulopathy in patients on platelet aggregation inhibitors immediately prior to renal transplantation with recombinant factor VIIa?

“…In our opinion, the effect of rFVIIa can be explained mainly by a disproportionately increased thrombin formation, which in turn leads to an enhanced inhibition of fibrinolysis via activation of thrombin activatable fibrinolysis inhibitor and also to an activation of platelets [13].…”