scholarly journals The endothelial protein C receptor plays an essential role in the maintenance of pregnancy

2020 ◽  
Vol 6 (45) ◽  
pp. eabb6196
Author(s):  
Michelle M. Castillo ◽  
Qiuhui Yang ◽  
Abril Solis Sigala ◽  
Dosia T. McKinney ◽  
Min Zhan ◽  
...  
Keyword(s):  
Pregnancy Outcomes ◽  
Protein C ◽  
Placental Abruption ◽  
Causal Link ◽  
Uterine Hemorrhage ◽  
Endothelial Protein C Receptor ◽  
Placental Development ◽  
Fetal Health ◽  
Placental Failure

Placenta-mediated pregnancy complications are a major challenge in the management of maternal-fetal health. Maternal thrombophilia is a suspected risk factor, but the role of thrombotic processes in these complications has remained unclear. Endothelial protein C receptor (EPCR) is an anticoagulant protein highly expressed in the placenta. EPCR autoantibodies and gene variants are associated with poor pregnancy outcomes. In mice, fetal EPCR deficiency results in placental failure and in utero death. We show that inhibition of molecules involved in thrombin generation or in the activation of maternal platelets allows placental development and embryonic survival. Nonetheless, placentae exhibit venous thrombosis in uteroplacental circulation associated with neonatal death. In contrast, maternal EPCR deficiency results in clinical and histological features of placental abruption and is ameliorated with concomitant Par4 deficiency. Our findings unveil a causal link between maternal thrombophilia, uterine hemorrhage, and placental abruption and identify Par4 as a potential target of therapeutic intervention.

Genetic Absence of Thrombin Receptor Par4 Overcomes the Obligate Requirement of EPCR in Mouse Placenta

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 425-425
Author(s):  
Michelle M Storage ◽  
Jianzhong An ◽  
Helena Liang ◽  
Qiuhui Yang ◽  
Mark Zogg ◽  
...  
Keyword(s):  
Protein C ◽  
Activated Protein C ◽  
Fold Increase ◽  
Thrombin Receptor ◽  
Mutant Form ◽  
Wild Type ◽  
Placental Development ◽  
Transgenic Expression ◽  
Placental Failure

Abstract Introduction: Murine models suggest that the Thrombomodulin-Protein C system plays a critical role in placentation and the maintenance of pregnancy. Severe Protein C deficiency in the mother results in pregnancy failure in early gestation. Thrombomodulin (Thbd) or the Endothelial Protein C Receptor (EPCR/ProcR) gene deletions result in embryonic death, secondary to developmental and functional abnormalities of the placenta. These molecules play multiple roles in coagulation and inflammation. The mechanisms governing their role in placental development and maintenance of placental function remain to be fully understood. The objective of this work is to identify the critical functions of EPCR and Thbd that are required for placental development. Both Thbd and EPCR augment activated protein C generation, albeit to different extents. We have examined if reduced activated Protein C generation mediates placental abnormalities of EPCR- and Thbd-null mice. Activation of thrombin receptors expressed on platelets and trophoblast cells can also contribute to placental failure. We examined the role of thrombin receptor Par4 in placental failure of EPCR-null mice. Methods: To assess the role of a PC generation in placental phenotype of Thbd- and EPCR-null mice, we used a transgene to express a hyperactivatable form of murine protein C (hMPC) under the control of transthyretin promoter. Thrombin cleaves this mutant form of Protein C 30-fold more efficiently than wild type protein C, without requiring the cofactor function of thrombomodulin. Wild type mice expressing hMPC show 2-fold increase in PC and 3-fold increase in aPC levels. hMPC expression in PC-null mice restores their ability to carry pregnancies. Breeding strategies were used to generate hMPCtg ProcR+/- or hMPCtg Thbd+/- female mice. These were mated to ProcR+/- or Thbd+/- males, respectively, and survival of ProcR-/- and Thbd-/- embryos was analyzed. Similar genetic strategy was used to analyze the role of thrombin receptor Par4 in the demise of EPCR-null embryos. Placental phenotypes and embryonic survival was compared with experiments in which the mother was continuously infused with LMWH using a subcutaneous osmotic pump. Results: As previously reported, EPCR-null mice die before 10.5 days post coitum (dpc) (ProcR+/- intercrosses, out of 41 live embryos none were ProcR-/-, 10 were expected, 21 aborted not genotyped, 7 pregnancies analyzed at 11.5 dpc) and none are found at wean (out of 30 live pups none were ProcR-/-, 8 were expected, 5 litters analyzed). Transgenic expression of hMPC in the mother resulted in some live ProcR-/- embryos at 11.5 dpc (4 ProcR-/- out of 41 live embryos, 10 were expected, 15 aborted not genotyped, 7 pregnancies at 11.5 dpc) and pups at wean (2 ProcR-/- out of 28 live, 7 litters analyzed). Despite transgenic hMPC expression ProcR-/- embryos and pups were underrepresented (P=0.007, chi square GOF test). Surviving ProcR-/- embryos showed normal placental histology grossly comparable to littermate controls. Expression of hMPC in the mother did not ameliorate fetal death of Thbd-null mice (out of 38 live embryos none were Thbd-/-, 10 expected, 16 aborted not genotyped, 7 pregnancies at 9.5 dpc). Continuous infusion of LMWH also resulted in some live ProcR-/- embryos at 11.5 dpc (3 ProcR-/- out of 19 live embryos, 5 expected, 11 aborted not genotyped, 3 pregnancies analyzed), but two were growth retarded and all 3 placentae showed markedly reduced placental labyrinth formation. In contrast to transgenic expression of hMPC and treatment with LMWH, when Par4-/- ProcR+/- animals were intercrossed, ProcR-/- animals were born at an expected Mendelian frequency (7 ProcR-/- out of 35 live pups, 9 expected, 7 litters analyzed). Conclusions: Our results show that transgenic expression of hMPC allows normal placental development and rescues a fraction of EPCR-null embryos. Thus, placental defect of EPCR-null mice is in part mediated by reduced generation of aPC on placental cells. In contrast to the transgenic expression of hMPC and LMWH treatment, genetic absence of Par4 completely overcame the placental defect and allowed development of EPCR-null embryos. Further studies will clarify contributions of maternal versus fetal Par4 in this phenomenon. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Endothelial Protein C Receptor plays an essential role in the maintenance of Pregnancy

2020 ◽  
Author(s):  
Michelle M Castillo ◽  
Qiuhui Yang ◽  
Abril Solis Sigala ◽  
Dosia T McKinney ◽  
Min Zhan ◽  
...  
Keyword(s):  
Factor Viii ◽  
Maternal Death ◽  
Pregnancy Outcomes ◽  
Pregnancy Complications ◽  
Protein C ◽  
Placental Abruption ◽  
Clotting Factor ◽  
List Type ◽  
Endothelial Protein C Receptor ◽  
Effect Relationship

ABSTRACTPlacenta-mediated pregnancy complications are a major challenge in the management of maternal-fetal health. Maternal thrombophilia is a suspected risk factor but the role of thrombotic processes in these complications and the potential for antithrombotic treatment have remained unclear. Endothelial Protein C Receptor (EPCR) is an anticoagulant protein highly expressed in the placenta. EPCR autoantibodies and specific gene variants of EPCR are associated with poor pregnancy outcomes. In mice, fetal EPCR deficiency results in placental failure and in utero death. Adult EPCR-deficient mice generated by maintaining placental expression exhibit plasma markers of thrombophilia without overt thrombosis. We demonstrate that inactivation of clotting factor VIII or Protease Activated Receptor 4 (Par4), Par3 or integrin αIIb in the mother allows placental development and intrauterine survival of murine embryos lacking EPCR. Rescued EPCR-deficient embryos exhibit thrombosis in placental venous sinuses at late gestation and a high rate of neonatal lethality. In contrast to fetal EPCR deficiency, maternal deficiency of EPCR results in frequent stillbirths and maternal death accompanied by pathological findings that resemble placental abruption and consumptive coagulopathy. Inactivation of Par4, but not clotting factor VIII, prevents maternal death and restores normal pregnancy outcomes. These observations establish a cause-effect relationship between maternal thrombophilia and placental abruption. They demonstrate that sites of uteroplacental thrombosis and the potential response to antithrombotic intervention may differ with gestational age and maternal versus fetal origin of thrombophilia. Our findings highlight the potential for therapeutic inhibition of thrombin-mediated platelet activation in a subset of pregnancy complications.KEY POINTSMurine model establishes a cause-effect relationship between maternal thrombophilia, retroplacental hemorrhage and severe pregnancy complications.Thrombin-mediated activation of maternal platelets is a key event in thrombophilia-associated pregnancy complications and a potential target of therapeutic intervention.Maternal venous channels in uteroplacental circulation are additional sites of thrombotic pathology associated with adverse neonatal outcomes.

Exploring the role of the phospholipid ligand in endothelial protein C receptor: A molecular dynamics study

2010 ◽  
Vol 78 (12) ◽  
pp. 2679-2690 ◽  
Author(s):  
Federica Chiappori ◽  
Ivan Merelli ◽  
Luciano Milanesi ◽  
Ermanna Rovida
Keyword(s):  
Molecular Dynamics ◽  
Protein C ◽  
Endothelial Protein C Receptor

scholarly journals Activation of endothelial intrinsic NF-κB pathway impairs protein C anticoagulation mechanism and promotes coagulation in endotoxemic mice

Blood ◽  
2009 ◽  
Vol 114 (12) ◽  
pp. 2521-2529 ◽  
Author(s):  
Dongmei Song ◽  
Xiaobing Ye ◽  
Honglei Xu ◽  
Shu Fang Liu
Keyword(s):  
Transgenic Mice ◽  
Tissue Factor ◽  
Protein C ◽  
Nuclear Factor Κb ◽  
Tissue Factor Expression ◽  
Wild Type ◽  
Endothelial Protein C Receptor ◽  
Lps Challenge ◽  
Factor Expression

Abstract Although the role of systemic activation of the nuclear factor κB (NF-κB) pathway in septic coagulation has been well documented, little is known about the contribution of endothelial-specific NF-κB signaling in this pathologic process. Here, we used transgenic mice that conditionally overexpress a mutant I-κBα, an inhibitor of NF-κB, selectively on endothelium, and their wild-type littermates to define the role of endothelial-specific NF-κB in septic coagulation. In wild-type mice, lipopolysaccharide (LPS) challenge (5 mg/kg intraperitoneally) caused markedly increased plasma markers of coagulation, decreased plasma fibrinogen level, and widespread tissue fibrin deposition, which were abrogated by endothelial NF-κB blockade in transgenic mice. Endothelial NF-κB blockade inhibited tissue factor expression in endothelial cells, but not in leukocytes. Endothelial NF-κB blockade did not inhibit LPS-induced tissue factor expression in heart, kidney, and liver. Endothelial NF-κB blockade prevented LPS down-regulation of endothelial protein C receptor (EPCR) and thrombomodulin protein expressions, inhibited tissue tumor necrosis factor-α converting enzyme activity, reduced EPCR shedding, and restored plasma protein C level. Our data demonstrate that endothelial intrinsic NF-κB signaling plays a pivotal role in septic coagulation and suggests a link between endothelial-specific NF-κB activation and the impairment of the thrombomodulin-protein C-EPCR anticoagulation pathway.

Laboratory Issues in Diagnosing Abnormalities of Protein C, Thrombomodulin, and Endothelial Cell Protein C Receptor

2002 ◽  
Vol 126 (11) ◽  
pp. 1337-1348 ◽  
Author(s):  
Kandice Kottke-Marchant ◽  
Philip Comp
Keyword(s):  
Endothelial Cell ◽  
Protein C ◽  
Medical Literature ◽  
Majority Vote ◽  
Diagnostic Testing ◽  
Laboratory Evaluation ◽  
Cell Protein ◽  
Endothelial Protein C Receptor ◽  
Protein C Deficiency

Abstract Objective.—To review the current understanding of the pathophysiology of protein C deficiency and its role in congenital thrombophilia. Recommendations for diagnostic testing for protein C function and concentration, derived from the medical literature and consensus opinions of recognized experts in the field, are included, specifying whom, how, and when to test. The role of related proteins, such as thrombomodulin and endothelial protein C receptor, is also reviewed. Data Sources.—Review of the published medical literature. Data Extraction and Synthesis.—A summary of the medical literature and proposed testing recommendations were prepared and presented at the College of American Pathologists Conference XXXVI: Diagnostic Issues in Thrombophilia. After discussion at the conference, consensus recommendations presented in this manuscript were accepted after a two-thirds majority vote by the participants. Conclusions.—Protein C deficiency is an uncommon genetic abnormality that may be a contributing cause of thrombophilia, often in conjunction with other genetic or acquired risk factors. When assay of protein C plasma levels is included in the laboratory evaluation of thrombophilia, a functional amidolytic protein C assay should be used for initial testing. The diagnosis of protein C deficiency should be established only after other acquired causes of protein C deficiency are excluded. A low protein C level should be confirmed with a subsequent assay on a new specimen. Antigenic protein C assays may be of benefit in subclassification of the type of protein C deficiency. The role of thrombomodulin and endothelial cell protein C receptor in thrombosis has yet to be clearly established, and diagnostic testing is not recommended at this time.

Placental beginnings: Reconfiguring placental development and pregnancy loss in feminist theory

2018 ◽  
Vol 20 (3) ◽  
pp. 283-298
Author(s):  
Sara DiCaglio
Keyword(s):  
Feminist Theory ◽  
Pregnancy Outcomes ◽  
Pregnancy Loss ◽  
Feminist Scholarship ◽  
Term Pregnancy ◽  
Placental Development ◽  
Hormone Production ◽  
Feminist Theories ◽  
Theories Of Subjectivity

The placenta has played an important role in feminist theories of subjectivity; however, the placenta of feminist theory has been the fully functional placenta of what is considered a successful full-term pregnancy. Pregnancy loss, a topic that has been generally overlooked within feminist scholarship, is absent from feminist theories of the placenta. This article uses early placental development, particularly development that takes place before the placenta becomes fully functional as an organ for hormone production and interchange, as a space through which to consider theorising subjectivity, reproduction and relation through pregnancy loss. In so doing, I argue that turning our attention to the placenta’s early development, regardless of outcomes, allows us to reimagine the role of process for feminist theories of subjectivity while also making room for a wider array of pregnancy outcomes, reinvigorating our ability to think about relations and models of hospitality that do not end as we might imagine.

scholarly journals Animal Models Demonstrate a Critical Role of Factor VIII in Par4- and Platelet-Mediated Pathology

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1057-1057
Author(s):  
Michelle Castillo ◽  
Quihui Yang ◽  
Dosia McKinney ◽  
Qizhen Shi ◽  
Rashmi Sood
Keyword(s):  
Embryonic Development ◽  
Platelet Activation ◽  
Protein C ◽  
Fetal Death ◽  
Hemophilia A ◽  
Adult Survival ◽  
Endothelial Protein C Receptor ◽  
Test Of Independence ◽  
Deficient Mice

Recent studies suggest that Tissue factor (TF)-initiated pathway can directly and rapidly yield FVIIIa-FIXa intrinsic tenase complex prior to coagulation amplification by thrombin (PMID 28729433). FVIII may also mediate TF/FVIIa-dependent FXa generation in the initiation phase of coagulation (PMID 30756344). These new insights prompted us to evaluate whether FVIII plays an important role in generating the minimum level of thrombin required for platelet activation in vivo. We used two distinct murine models of platelet- and Par4-mediated pathology to address this question. We have previously shown that mice homozygous for a hypomorphic thrombomodulin allele (ThbdPro/Pro) are aborted in pregnancies of FV Leiden (FVQ/Q) females before day 12.5 of gestation (Table 1, row 1; PMID 23325830). Par4-activation on maternal platelets plays a key role in fetal death in this model; its absence allows embryonic development and restores Mendelian frequency of live ThbdPro/Pro embryos found at term (Table 1, row 2; PMID 17438064). We examined the effect of reduced TF or genetic deletion of FVIII on embryonic survival of ThbdPro/Pro embryos. We demonstrate that inhibition of TF or the absence of FVIII, each significantly protect the fetus from maternal platelet-mediated death (Table 1, rows 3 & 4). Treatment of the mother with anti-TF antibody 1H1 resulted in 11 ThbdPro/Pro embryos (30 analyzed, 50% or 15 expected; 37% observed, 95% CI 20 to 56%). In the absence of FVIII, 10 ThbdPro/Pro embryos were observed (30 analyzed, 50% or 15 expected; 37% observed, 95% CI 20 to 56%). The outcome of TF inhibition was comparable to the absence of FVIII (P=0.79 Χ2 test of independence). In a second model, Endothelial Protein C Receptor null mice (ProcR-/-) die in midgestation (Table 2, row 1), but reducing TF expression restores Mendelian frequency of ProcR-/- pups (PMID 15956290). We find that ProcR-/- embryos survive past midgestation if the mother is injected with platelet depleting antibodies. Genetic absence of Par4 allows ProcR-/- mice to survive embryonic development (Table 2, row 2; 25% or 18 ProcR-/- neonates expected, 13 observed, 95% CI 10.3 to 29.7%). We investigated the role of FVIII in the midgestational demise of ProcR-/- mice. We demonstrate that genetic absence of FVIII also protects ProcR-/- mice from intrauterine demise (Table 2, row 3; 25% or 12 out of 48 ProcR-/- expected, 12 observed, 95% CI 13.6 to 29.6%). Similar to TF reduction, the absence of FVIII restores Mendelian frequency of ProcR-/- embryos at term. Thus, the outcome from the genetic absence of FVIII was comparable to the absence of Par4 (P=0.4 Χ2 test of independence). In a third model, we have established that epiblast restricted deletion results in normal Mendelian frequency of thrombomodulin-deficient embryos (Meox2Cretg Thbdδ/δ) at term, but about 1/3rd survive the trauma of birth. Most surviving mice later succumb to severe tail and limb thrombosis and several exhibit circling behavior indicative of stroke. Genetic absence of Par4 does not improve perinatal survival, but protects thrombomodulin-deficient mice from stroke and thrombotic disease, significantly improving adult survival. We tested whether genetic absence of FVIII affords protection similar to the absence of Par4. We report that similar to Par4-/- Meox2Cretg Thbdδ/δ mice, FVIII-/- Meox2Cretg Thbdδ/δ mice do not exhibit thrombosis or stroke (current age range 2 to 8 months). Thus, the absence of FVIII is protective in Par4-mediated thrombosis and stroke observed in thrombomodulin-deficient mice. These studies establish an essential role of FVIII in Par4-mediated pathologies. They strongly support the notion that FVIII activation is required for sufficient accumulation of thrombin for platelet activation. Notably, these are models of physiological thrombosis or platelet-mediated fetal death, without the use of artificial injury or insult. All three models that we describe lack a fully functional system for generating activated protein C (aPC). Our results suggest that inactivation of FVIII by aPC may be necessary to tightly regulate thrombin-mediated platelet activation. Reduced platelet activation observed in hemophilia A mice, albeit in the context of prevention of thrombosis, also lends support to the notion that defects in primary hemostasis due to reduced platelet activity could be an important component of bleeding diathesis in hemophilia A patients. Disclosures No relevant conflicts of interest to declare.

The endothelial protein C receptor impairs the antibacterial response in murine pneumococcal pneumonia and sepsis

2014 ◽  
Vol 111 (05) ◽  
pp. 970-980 ◽  
Author(s):  
J. Daan de Boer ◽  
Liesbeth M. Kager ◽  
Joris J. T. H. Roelofs ◽  
Joost C. M. Meijers ◽  
Charles T. Esmon ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Bacterial Growth ◽  
Protein C ◽  
Pneumococcal Pneumonia ◽  
Wild Type ◽  
Endothelial Protein C Receptor ◽  
Pulmonary Endothelium ◽  
Pneumococcal Sepsis ◽  
Bacterial Outgrowth

SummaryPneumococcal pneumonia is a frequent cause of gram-positive sepsis and has a high mortality. The endothelial protein C receptor (EPCR) has been implicated in both the activation of protein C (PC) and the anti-inflammatory actions of activated (A)PC. The aim of this study was to determine the role of the EPCR in murine pneumococcal pneumonia and sepsis. Wild-type (WT), EPCR knockout (KO) and Tie2-EPCR mice, which overexpress EPCR on the endothelium, were infected intranasally (pneumonia) or intravenously (sepsis) with viable Streptococcus pneumoniae and euthanised at 24 or 48 hours after initiation of the infection for analyses. Pneumonia did not alter constitutive EPCR expression on pulmonary endothelium but was associated with an influx of EPCR positive neutrophils into lung tissue. In pneumococcal pneumonia EPCR KO mice demonstrated diminished bacterial growth in the lungs and dissemination to spleen and liver, reduced neutrophil recruitment to the lungs and a mitigated inflammatory response. Moreover, EPCR KO mice displayed enhanced activation of coagulation in the early phase of disease. Correspondingly, in pneumococcal sepsis EPCR KO mice showed reduced bacterial growth in lung and liver and attenuated cytokine release. Conversely, EPCR-overexpressing mice displayed higher bacterial outgrowth in lung, blood, spleen and liver in pneumococcal sepsis. In conclusion, EPCR impairs antibacterial defense in both pneumococcal pneumonia and sepsis, which is associated with an enhanced pro-inflammatory response.

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