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The effects of flow on the activation of bovine prothrombin by prothrombinase at physiologically relevant shear rates

The generation of thrombin by preassembled prothrombinase on phospholipid coated capillaries was studied under laminar flow at physiologic shear rates (100–1000 sec−1). When prothrombin (1.4μM) was perfused, thrombin levels reached a steady‐state that decreased with increasing shear rate; however, generation was independent of shear rate when corrected for the velocity of the effluent. The ratio of α‐thrombin to meizothrombin formed was 3:2 at shear rates of 250 and 500 sec−1. Kinetic constants determined at a shear rate of 250 sec−1 were in agreement with those obtained in closed systems, suggesting that the exchange between the bulk solution and the capillary wall region is limited by the competition between molecular diffusion of thrombin and flow convection. This results in the development of a diffusive boundary layer that spatially confines thrombin, yielding predicted concentrations of up to 1μM. The observation of extensive thrombin feedback cleavage of the phospholipid binding domain from prothrombin and meizothrombin is consistent with such high concentrations of thrombin. A flow transport model is presented for thrombin generation that estimates the development of the thrombin layer. Supported by NIH HL46703 and 5T32HL007594.