Extraembryonic expression of EPCR is essential for embryonic viability

Abstract

The endothelial cell protein C receptor (EPCR) augments protein C activation by the thrombin-thrombomodulin complex. Deletion of the EPCR gene (Procr) in mice leads to embryonic lethality before embryonic day 10 (E10.0). EPCR is detected in the giant trophoblast cells at the fetomaternal boundary from E7.5 and weakly in embryonic aortic endothelial cells from E13.5, suggesting that extraembryonic EPCR expression may be essential for embryonic viability. Using conditional knock-out strategies, we demonstrate that Procr-deficient embryos with EPCR expression on placenta giant trophoblasts can be carried to term and then develop normally. Conversely, EPCR expression in the embryo, without expression in the giant trophoblast cells, does not rescue the mice. In genetically modified mice with low tissue factor activity, Procr deficiency is not lethal to the embryo. As adults, Procr-deficient mice generate more thrombin and activate less protein C in response to procoagulant stimuli. Spontaneous thrombin formation in the deficient animals increases with age. These findings show that extraembryonic EPCR expression is critical for embryo development. IntroductionThe protein C anticoagulant pathway serves as an "on-demand" anticoagulant system that controls thrombin generation by inhibiting factors Va and VIIIa. 1 Thrombin not only activates platelets and clots fibrinogen, but also binds to thrombomodulin (TM) on the endothelium, at which time these procoagulant activities of thrombin are blocked, while conversion of protein C to the anticoagulant serine proteinase, activated protein C (APC), is augmented. 2 Endothelial cell protein C receptor (EPCR) binds protein C and increases the rate of protein C activation by the thrombin-TM complex in cell culture 3 and in vivo. 4 EPCR overexpression on the endothelial cell surface increases the rate of protein C activation further. 5 Inhibition of EPCR-protein C interaction converts the response to sublethal concentrations of Escherichia coli into a lethal response. These animals exhibit disseminated intravascular coagulation, intense neutrophil influx into the tissues, and elevation of some of the inflammatory cytokines. 6 In adults, EPCR is localized primarily on endothelial cells of large blood vessels and is very low or absent from the microvascular endothelium of most tissues. 7 EPCR is also abundant on the trophoblast giant cells at the feto-maternal boundary from embryonic day 7.5 (E7.5), suggesting a role in the hemostatic regulation of the maternal blood that irrigates these surfaces. Deletion of the EPCR gene (Procr) in mice leads to embryonic lethality before E10.0. (Gene symbols used in this paper are in accordance with the style guidelines of the National Center for Biotechnology Information, National Institutes of Health. Specifically, the gene encoding EPCR is designated Procr; tissue factor, F3; thrombomodulin, Thbd; Tie2, Tek; and Mox2, Meox2.) At this time, excess fibrin deposition is detected around the trophoblast giant cells (derived from the Procr Ϫ/Ϫ embryo...

Endothelial Cell Protein C Receptor Gene 6936a/G, 1651c/G, 4678g/C Polymorphisms and Soluble Endothelial Protein C Receptor Levels Impact on in Vitro Fertilization Outcomes

“…Soluble EPCR plays a role in maintenance of pregnancy. That is confirmed by the findings that EPCR expression is critical for embryo development [11]. In addition, blocking of EPCR activity by anti-EPCR autoantibodies was associated with adverse pregnancy outcomes [12].…”

“…EPCR expression is essential for embryo development, this is supported by the accelerated embryonic lethality observed in EPCR knockout mice before embryonic day 10. This is owing to the fact that fibrin deposition around trophoblast giant cells results in thrombosis at the maternal-embryonic interface [11]. In addition, EPCR activity blockade by anti-EPCR autoantibodies was associated with adverse pregnancy outcomes [12].…”

Maternal Par4 and platelets contribute to defective placenta formation in mouse embryos lacking thrombomodulin

“…14 However, in striking contrast to endothelial cells, trophoblast cells constitutively express the initiator of coagulation, tissue factor (TF), 14,[17][18][19][20] and genetic suppression of TF activity on trophoblast cells restores normal development of mice lacking Thbd or EPCR. 16,17 This suggests that the predominant function of Thbd and EPCR on trophoblast cells is to provide an anticoagulant mechanism, which counteracts unfettered, TF-initiated coagulation activation in the placenta. Additional evidence for APC-mediated anticoagulation as the critical Thbd function on trophoblast cells stems from the observation that maternal APC resistance secondary to the Leiden mutation in coagulation fV, when combined with reduced Thbd function on trophoblast cells secondary to fetal Thbd-Pro mutation, replicates the phenotype of Thbd-null embryos.…”

“…12,13 These give rise to the bulk of the fetal-derived cell mass of the placenta, and fulfill an endothelial cell-like role in the regulation of hemostasis in the placental vascular bed. 14 Cell type-selective restoration of Thbd or EPCR expression in trophoblast cells restores normal placental development of Thbd-and EPCR-knockout embryos, respectively, 15,16 demonstrating that the critical cell type on which either receptor must be expressed is not the vascular endothelium, but trophoblast cells. A loss of Thbd function is associated with a failure of trophoblast cells to maintain proliferation in the ectoplacental cone and gives rise to a population of differentiated derivatives that establish the bulk of the mature placenta.…”

Consultative Hematology: The Pregnant Patient Pregnancy Loss

“…Structural abnormalities (thinning of the layer lining the maternal lacunae and reduced number of trophoblast cellular contacts) are seen in the placentae of tissue-factor null mice embryos, 7 whereas absence of thrombomodulin is associated with mice fetal death that is not associated with fibrin deposition, suggesting a responsible mechanism other than thrombosis. 8 Inactivation of the gene for protein C 9 and endothelial protein C receptor gene deletion 10 are also associated with mice embryo death. In vitro observations suggest that the presence of activated coagulation factors results in cell-type specific changes in trophoblast gene expression.…”