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        Studies on Coagulation Incompatibilities for Xenotransplantation

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        Microvascular thrombosis, following the activation of clotting cascade, is a hallmark of porcine solid organ xenograft rejection. The analysis of differences between human, monkey, and pig coagulation systems is crucial when monkey is used as animal model and pig as organ donor in xenotransplantation. Thrombosis, according to many authors, may be due to the molecular incompatibilities between natural anticoagulants present on pig endothelium and primate activated coagulation factors. The generation of activated protein C (PC) is critical for the physiological anticoagulation. One of the major incompatibilities may be related to the inability of pig thrombomodulin (TM) and endothelial protein C receptor to activate the recipient (primate) circulating PC in the presence of thrombin. Tissue factor pathway inhibitor (TFPI), is the primary inhibitor of tissue factor (TF)-induced coagulation. TFPI directly inhibits the activated factor X (FXa) and blocks the procoagulant activity of the TF/factor VIIa (FVIIa) complex by forming a quaternary TF/FVIIa/FXa/TFPI complex. Microvascular thrombosis, observed in the organ transplant, may also be due to the failure of pig TFPI to bind human FXa efficiently and inhibit human FVIIa/TF activity. The methods described in this chapter can be useful for the identification and characterization of primate and pig coagulation factors (isolated from a small volume of blood) by using SDS-PAGE and immunoblotting. Differences in molecular weight can help in the identification of the origin (pig or primate) of coagulation proteins in plasma from the recipient of xenografts. On the other hand, in vitro models of PC pathway and TFPI on human umbilical vein endothelial cells (HUVEC) and porcine aortic endothelial cells (PAEC) are described which can be used for studying incompatibilities between primate and pig.
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