thromboplastin and Abortion--Spontaneous

The paternal antigens presented by the fetus could be considered foreign by the mother's immune system and elicit an immune response. Here we show that the complement system functions as an effector in fetal rejection in two different mouse models of pregnancy loss. In a mouse model of fetal loss and growth restriction (IUGR) induced by antiphospholipid antibodies (aPL), we found that complement activation is a crucial and early mediator of pregnancy loss. We demonstrated that C5a-C5aR interaction and neutrophils are key mediators of fetal injury. We identified tissue factor (TF) as a critical intermediate that, acting downstream of C5 activation, enhances neutrophil activity and trophoblast injury. In an antibody-independent mouse model of spontaneous miscarriage and IUGR (CBAxDBA) we also identified C5a as an essential mediator. Complement activation caused dysregulation of the angiogenic factors (deficiency of free vascular endothelial growth factor (VEGF) and elevated levels of soluble VEGF receptor 1) required for normal placental development. Inhibition of complement activation prevented angiogenesis failure and rescued pregnancies. Our studies in antibody-dependent and antibody-independent models of pregnancy complications identified complement activation as the crucial mediator of damage and will allow development of new interventions to prevent pregnancy loss and IUGR.

Topics: Abortion, Spontaneous; Animals; Antibodies, Antiphospholipid; Anticoagulants; Complement Activation; Complement C5a; Complement Inactivating Agents; Female; Fetal Death; Fetal Growth Retardation; Growth Inhibitors; Humans; Macrophages; Mice; Neutrophils; Oxidative Stress; Pregnancy; Receptor, Anaphylatoxin C5a; Thromboplastin; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-1

The spontaneous loss of normal karyotype embryos may be initiated or prevented by the maternal immune system. In mice, loss between the time of implantation (day 4.5) and formation of a vascularized placenta (day 9.5) when the embryo is too large to survive by diffusion alone, is analogous to occult pregnancy failure in humans. They are called occult because usually the woman does not know she is pregnant. From studies in mice, these early losses have a different mechanism than abortion of a vascularized placenta (analogous to clinically evident human spontaneous miscarriage). The latter depend on the activation of the novel prothrombinase fgl2 on the fetal trophoblast and in maternal decidua by the T helper-1 (Th1) type cytokines TNF- alpha+gamma -interferon that arise from NK cells and NK gammadelta T cells; conversion of prothrombin to thrombin which in turn generates IL8 that activates polymorphonuclear leukocytes leads to embryonic death. These inflammatory processes are counteracted by Th2/3-type cytokines that arise in part from V gamma 1 delta 6 T cells reacting to, as yet, unidentified trophoblast antigens in the presence of the 'tolerance signaling molecule' OX-2. By contrast, peri-implantation losses (between implantation and formation of a vascularized placenta, analogous to occult losses in humans) appear to be dependent upon perforin(+)cells, complement activation, and products of alphabeta T and NK alphabeta T cells, but not on TNF- alpha or procoagulant activation. Similarities and differences between findings in the mouse and human, and the potential evolutionary significance of mechanisms affecting reproductive success are reviewed.

Topics: Abortion, Spontaneous; Animals; Biological Evolution; Embryo Implantation; Female; Fibrinogen; Humans; Immune Tolerance; Maternal-Fetal Exchange; Mice; Pregnancy; Thromboplastin; Trophoblasts

Antiphospholipid syndrome (APS) is a systemic autoimmune disorder of young adults associated with devastating pregnancy complications (recurrent miscarriages, preeclampsia and low birth weight) and vascular complications including thrombosis. The key components implicated in pathogenesis of APS are the complement cascade and tissue factor (TF) activity causing inflammation and coagulation. Purinergic signalling involving catabolism of ATP to adenosine by cell-surface enzymes CD39 and CD73 has anti-inflammatory and anti-thrombotic effects. We studied whether activities of CD39 and CD73 are important in preventing the development of miscarriages in APS.. We studied frequency of miscarriages and decidual pathology following passive transfer of human aPL-ab to pregnant wildtype mice, and mice deficient in CD39 and CD73, and also transgenic mice exhibiting 2-3X higher CD39 activity.. aPL-ab infusion in pregnant CD39-or CD73-knockout mice triggers an increase in miscarriages, associated with increased TF expression and complement deposition as well as elevated oxidative stress and pro-inflammatory TNF-α and IL-10 expression within the placental decidua. In contrast, aPL-ab induced miscarriages are prevented in mice over-expressing CD39, with reduced decidual TF expression and C3d deposition, diminished lipid peroxidation (4-hydroxynonenal or 4-HNE positive lipid adducts), and reduced TNF-α expression.. We demonstrate a protective role for CD39 in APS and provide rationale for both the development of endothelial cell-targeted soluble CD39 as a novel therapeutic for APS and analysis of perturbations in the purinergic pathway to explain human disease.

Topics: 5'-Nucleotidase; Abortion, Spontaneous; Adult; Animals; Antibodies, Antiphospholipid; Antigens, CD; Antiphospholipid Syndrome; Apyrase; Complement C3d; Disease Models, Animal; Female; Humans; Immunization, Passive; Inflammation; Inflammation Mediators; Lipid Peroxidation; Mice; Mice, Knockout; Mice, Transgenic; Pregnancy; Pregnancy Complications; Thromboplastin; Tumor Necrosis Factor-alpha

Pregnancy loss and intrauterine growth restriction (IUGR) are serious pregnancy complications, and the triggers and mediators of placental and fetal damage are not completely understood. Using a mouse model of recurrent spontaneous miscarriages (DBA/2-mated CBA/J mice) that shares features with human recurrent miscarriage and fetal growth restriction, we identified tissue factor (TF) as an essential participating factor in placental and fetal injury. We have previously shown that C5a releases antiangiogenic molecule sFlt-1 in monocytes that causes defective placental development and fetal death in DBA/2-mated CBA/J mice. In this study, we found that TF not only activates the coagulation pathway, but it also mediates sFlt-1 release in monocytes causing defective placental development and fetal death. Blockade of TF with a monoclonal antibody inhibited sFlt-1 release, prevented the pathological activation of the coagulation pathway, restored placental blood flow, prevented placental oxidative stress, and rescued pregnancies. We also demonstrated that pravastatin, by down-regulating TF expression on monocytes and trophoblasts, prevented placental damage and protected pregnancies in DBA/2-mated CBA/J mice. These studies indicate that TF is an important mediator in fetal death and growth restriction and that statins may be a good treatment for women with recurrent miscarriages and IUGR.

Topics: Abortion, Spontaneous; Animals; Animals, Newborn; Anticoagulants; Antithrombin III; Cells, Cultured; Disease Models, Animal; Female; Humans; Infant, Newborn; Male; Mice; Monocytes; Nitrogen Oxides; Oxidative Stress; Placenta; Pravastatin; Pregnancy; Protein Binding; Thrombin; Thromboplastin; Trophoblasts; Vascular Endothelial Growth Factor Receptor-1

Disruption of the mouse gene encoding the blood coagulation inhibitor thrombomodulin (Thbd) leads to embryonic lethality caused by an unknown defect in the placenta. We show that the abortion of thrombomodulin-deficient embryos is caused by tissue factor-initiated activation of the blood coagulation cascade at the feto-maternal interface. Activated coagulation factors induce cell death and growth inhibition of placental trophoblast cells by two distinct mechanisms. The death of giant trophoblast cells is caused by conversion of the thrombin substrate fibrinogen to fibrin and subsequent formation of fibrin degradation products. In contrast, the growth arrest of trophoblast cells is not mediated by fibrin, but is a likely result of engagement of protease-activated receptors (PAR)-2 and PAR-4 by coagulation factors. These findings show a new function for the thrombomodulin-protein C system in controlling the growth and survival of trophoblast cells in the placenta. This function is essential for the maintenance of pregnancy.

Topics: Abortion, Spontaneous; Animals; Blood Coagulation; Cell Division; Embryo Loss; Embryo, Mammalian; Female; Fibrin; Fibrinogen; Fibrinolysin; In Situ Hybridization; In Situ Nick-End Labeling; Male; Mice; Mice, Inbred C57BL; Placenta; Pregnancy; Pregnancy Maintenance; Protein C; Receptor, PAR-2; Receptors, Thrombin; Thrombin; Thrombomodulin; Thromboplastin; Trophoblasts

The immune system contributes to the outcome of pregnancy by complex immunological interactions. Cytokines especially influence the immune milieu pro or contra pregnancy. T helper 1 (Th1) cytokines [tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma)] cause inflammation and together are thought to threaten the maintenance of pregnancy. It has been proposed that increased levels of these Th1 cytokines activate coagulation via up-regulating the novel prothrombinase, fgl2. This study further investigates the Th1 cytokine up-regulation of fgl2 expression in a pathophysiological, stress induced abortion model, and an inflammatory, interleukin-12 (IL-12) triggered abortion model.. The DBA/2J-mated CBA/J female mice were exposed to sonic sound stress or were injected with IL-12 during early gestation. On day 13.5 of pregnancy the uteri were removed and the resorption rate was calculated. We evaluated TNF-alpha, IFN-gamma, fgl2 as well as IL-12 messenger RNA (mRNA) expression in decidual samples of all mice by quantitative, real-time polymerase chain reaction (PCR).. A similar resorption rate of 24% was detected in stressed mice, as well as in IL-12 injected mice compared with approximately 11% in non-stressed, non-injected control mice. In stressed mice compared with controls, we observed on day 13.5 up-regulated TNF-alpha, unchanged IFN-gamma down-regulated fgl2, and a slightly increased levels of IL-12. In the IL-12 triggered abortion model, we observed up-regulated levels of TNF-alpha, IFN-gamma and fgl2.. These novel data suggest two distinct cytokine patterns leading to similar abortion rates. A physiological cascade associated with up-regulation of TNF-alpha, and an IL-12-triggered cascade characterized by persistent up-regulation of TNF-alpha and IFN-gamma as well as a persistent increase in fgl2.

Topics: Abortion, Spontaneous; Adjuvants, Immunologic; Animals; Female; Inflammation; Interferon-gamma; Interleukin-12; Mice; Models, Animal; Pregnancy; Reverse Transcriptase Polymerase Chain Reaction; Stress, Psychological; Thromboplastin; Tumor Necrosis Factor-alpha; Up-Regulation

Clinicians commonly evaluate patients with thrombosis or a prolonged activated partial thromboplastin time (aPTT) for the presence of a lupus anticoagulant (LA). We evaluated strategies for detecting LA, in three clinical settings, with decision-modeling techniques. A decision tree was constructed with 12 strategies, using a combination of aPTT and dilute Russell viper venom times (dRVVT) with confirmatory tests, tissue thromboplastin time (TTI), platelet neutralization procedures, and mixing studies. Probabilities and costs of adverse events and test costs were obtained from the literature. Patient preference for each strategy was evaluated by assigning utilities to each outcome. On the basis of assay results in 90 healthy people and 77 patients, we calculated sensitivities and specificities for each strategy, with true positives defined as suggested by the International Society on Thrombosis and Haemostasis. The least costly strategy for evaluation of patients with a prolonged aPTT, or with thrombosis, is not to test and to assume that LA is absent. For patients with systemic lupus erythematosus (SLE), it is least expensive not to test, although testing with TTI alone can also be considered an efficient strategy. The strategy of highest utility to patients with SLE is testing with TTI, followed by dRVVT. On the basis of these cost and utility results, clinicians' strategies for detecting LA may need modification. These strategies would then optimally be tested in clinical trials.

Topics: Abortion, Spontaneous; Autoantibodies; Costs and Cost Analysis; Decision Trees; Humans; Lupus Coagulation Inhibitor; Lupus Erythematosus, Systemic; Partial Thromboplastin Time; Prothrombin Time; Sensitivity and Specificity; Thromboplastin; Thrombosis

Spontaneous abortion of normal karyotype embryos in mice and in humans is associated with an increase in uterine T helper (Th) 1 type proinflammatory cytokines, tumour necrosis factor (TNF)-alpha, interferon-gamma and interleukin (IL)-1, and a deficiency of Th2/3 type cytokines, IL-4, IL-10, and transforming growth factor (TGF)-beta2. In mice, Th1 cytokines up-regulate a novel prothrombinase, fgl2, which via thrombin, leads to activation of polymorphonuclear leukocytes that terminate the pregnancy. Here we show that Th1 cytokines up-regulate fgl2 mRNA in fetal trophoblast and secondary decidua of CBA/JxDBA/2 and CBA/JxBALB/c matings, and promote fibrin deposition. This pattern is accompanied by a high rate of abortion. However, the spontaneous abortion rates in abortion-prone CBAxDBA/2 matings and in low abortion rate CBAxBALB/c matings were significantly lower than that expected from the frequency of implantations with high levels of fibrin and fgl2 mRNA(hi). As the glycoprotein OX-2 occurs in the pregnant rat uterus and can deviate cytokine responses to Th2/3, we investigated OX-2 in pregnant CBA/J mice. We found OX-2 mRNA was present at the same sites as fgl2 mRNA, but was reduced in response to Th1 cytokines. Furthermore, anti-OX-2 raised the abortion rate to predicted levels, while recombinant OX-2 dramatically reduced the abortion rate. Fgl2 prothrombinase may provide a mechanism explaining pregnancy loss, and conversely, successful pregnancy may be due in part to OX-2-dependent activation of maternal tolerance mechanisms at the feto-maternal interface.

Topics: Abortion, Spontaneous; Animals; Antigens, CD; Antigens, Surface; Base Sequence; Cytokines; Decidua; DNA Primers; Female; Fibrin; Fibrinogen; Gene Expression; Humans; In Situ Hybridization; Male; Mice; Mice, Inbred BALB C; Mice, Inbred CBA; Mice, Inbred DBA; Pregnancy; RNA, Messenger; Thromboplastin; Trophoblasts

Impaired trophoblast invasion during the first trimester of pregnancy is linked to spontaneous abortion, and defective invasion in the second trimester to hypertension + proteinuria (pre-eclampsia). Hypertension developing during the third trimester of human pregnancy represents, in part, a corrective response in the mother to provide adequate placental perfusion for fetal growth when trophoblast has not to invaded and converted the myometrial porprtion of maternal spiral arteries into to low resistance-high capacitance conduits. Deportation of vesicles from hypoxemic trophoblast is thought to cause hypertension plus proteinuria, vascular damage and a systemic coagulopathy. Trophoblast invasion may be inhibited by local cytokines, such as TGF-betas but Thl-type cytokines associated with pre-eclapmsia and spontaneous abortions (e.g., IL-1, TNF-alpha, IFN-gamma) are not known to inhibit migration at in situ concentrations. Trophoblast invasion is also inhibited by the binding of surface integrins to fibronectin and fibrin, and fibrin production is stimulated by these Th1 cytokines via up-regulation of prothrombinases(s) such as fg12 which directly and via TNF-alpha-facilitated inflamation compromise trophoblast cell integrity. We, therefore, asked if fg12 expression and TNF-alpha are increased in first trimester human miscarriage and in third trimester pre-eclampsia.. fg12 mRNA was detected using in situ hybridization and fg12 protein by immunohistochemistry. TNF-alpha mRNA and protein were similarly tested. The techniques were validated using uterine sections from day 8.5 of CBA x DBA/2 pregnancies, and then were applied to sections of placentae from normal and pre-eclamptic pregnancies with and without intrauterine fetal growth restriction (IUGR). Fibrin was detectectd by immunohistochemistry.. Expression of fg12 protein correlated with fg12 mRNA expression in mouse uteri and in placentae from normal human pregnancies. Increased expression of fg12 and TNF-alpha mRNA and protein, and increased fibrin deposition was detected in placental trophoblast.. Activation of fg12 prothrombinase by Th1-type cytokines in pregnancy may lead to spontaneous abortion, or in ongoing pregnancy, to pre-eclampsia and/or IUGR.

Topics: Abortion, Spontaneous; Animals; Enzyme Activation; Female; Humans; In Situ Hybridization; Male; Mice; Mice, Inbred BALB C; Mice, Inbred CBA; Mice, Inbred DBA; Placenta; Pre-Eclampsia; Pregnancy; Pregnancy Trimester, First; Pregnancy Trimester, Third; RNA, Messenger; Thromboplastin; Tumor Necrosis Factor-alpha

Abortion of chromosomally normal embryos in the CBA X DBA/2 mating combination is triggered by release of Th1 cytokines (tumor necrosis factor [TNF]-alpha, interferon [IFN]-gamma, and interleukin [IL]-1), which cause abortion via a novel prothrombinase, Fgl2, and polymorphonuclear leukocytes. The site of activation may be maternal vascular endothelium on arteries and veins nourishing the placenta. Activation of coagulation is also prominent in spontaneous abortion of chromosomally normal human embryos. We asked where is Fgl2 up-regulated in the uterus in murine abortions, and if similar Fgl2 expression occurs in human pregnancy failure.. Control CBA X DBA/2 pregnant mice, or from mice injected with TNF-alpha + IFN-gamma on day 7.5 of gestation, were removed on day 8.5, fixed, sectioned, and subject to in situ hybridization for Fgl2. Sections were also stained for fibrin. Elective first trimester termination samples or biopsies taken early in the course of a recurrent miscarriage were similarly fixed, sectioned, and analyzed by in situ hybridization. Control and cytokine-treated mice were anticoagulated with heparin, an activator of antithrombin III, and/or the direct anti-thrombin inhibitor hirudin.. Low level Fgl2 expression localized to basal decidua remote from the embryo was noted in control mice; cytokine treatment, which causes greater than 80% of abortions, produced a striking up-regulation in this area as well as in a band at the junction of decidua and myometrium. Trophoblast also became strikingly positive. Fgl2 expression was associated with increased fibrin staining. Anticoagulation significantly protected against abortions, but doses were limited by the complication of retroplacental hemorrhage. In tissue from normal first trimester pregnancy, minimal Fgl2 positivity was seen in some villous syncytiotrophoblast, in villous stroma, cytotrophoblast, and in some cells in decidua. In spontaneous abortion of normal embryo, striking Fgl2 positivity was seen in syncytiotrophoblast and extravillous cytotrophoblast, in association with areas of thrombus formation.. Fgl2 appears to be physiologically expressed and may protect against the internal danger of maternal and/or fetal bleeding during pregnancy and at parturition; a role in inhibiting transplacental traffic is also possible. External dangers in the form of stress, endotoxin, and antigens eliciting Th1 cytokine responses upregulate Fgl2 prothrombinase in trophoblast as well as in decidua, which results in spontaneous abortion of immunogenetically "weaker" embryos.

Topics: Abortion, Spontaneous; Animals; Antithrombins; Decidua; Female; Fibrinogen; Heparin; Hirudins; Humans; Interferon-gamma; Male; Mice; Mice, Inbred CBA; Mice, Inbred DBA; Pregnancy; Pregnancy Trimester, First; Thromboplastin; Trophoblasts; Tumor Necrosis Factor-alpha

Spontaneous resorption in the CBA x DBA/2 model is attributed to NK cells, macrophages, and Th1-type cytokines. In vivo depletion of NK cells by anti-asialoGM1 Ab or macrophage depletion by silicon dioxide treatment reduced abortion rates, which could no longer be boosted by injecting TNF-alpha (which activates NK cells) or IFN-gamma (which activates macrophages). TNF-alpha + gamma-IFN coadministration aborted >80% of the embryos whether or not NK cells or macrophages had been depleted or estradiol + progesterone was injected to correct potential reduction in ovarian function by cytokines. The cytokines also aborted IRF1+/+ C57BL/6 but not IRF1-/- females pregnant by IRF1+/+ DBA/2. Both spontaneous and cytokine-boosted abortions in CBA x DBA/2 were blocked by Ab to fgl2 prothrombinase [corrected] expressed by cytokine-stimulated vascular endothelial cells and monocytes; in vivo Ab depletion of granulocytes also prevented TNF-alpha + IFN-gamma-induced abortions. Cytokine-triggered thrombotic/inflammatory processes in maternal uteroplacental blood vessels causes abortion.

Topics: Abortion, Spontaneous; Animals; Crosses, Genetic; Cytokines; DNA-Binding Proteins; Endothelium, Vascular; Female; Fibrinogen; Granulocytes; Immune Sera; Injections, Intraperitoneal; Interferon Regulatory Factor-1; Interferon-gamma; Killer Cells, Natural; Macrophages; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Inbred DBA; Mice, Knockout; Phosphoproteins; Pregnancy; Thromboplastin

The clinical and laboratory findings in seven female patients with primary autoimmune diseases, one female patient with lymphoplasmacytoid (LP) immunocytoma and IgM paraproteinemia, and two male patients with multiple myeloma are described. The common denominator in all patients was a lupus anticoagulant or a closely related coagulation disorder. Recurrent thrombosis was observed in six patients with autoimmune diseases and in two patients with malignant monoclonal gammopathies. Other clinical manifestations included cerebral disorders (four patients with autoimmune disease/two patients with monoclonal gammopathy), repeated obstetric complications (6/1), asymptomatic valvular heart disease (6/1), renal dysfunction (6/2), hepatic involvement (2/2), and arthropathy (2/0). Laboratory investigations revealed a biologic false-positive serological test for syphilis in six patients with autoimmune disease and one with monoclonal gammopathy, antinuclear antibodies (4/0), antibodies against DNA (4/1), and a positive direct Coombs test (3/1) which was accompanied by hemolytic anemia in two patients (1/1). Additionally slight leukocytopenia (2/1) and thrombocytopenia (6/2) were observed; abnormal bleeding was only seen in one patient with severe thrombocytopenia. Other complications characteristic of LP immunocytoma or multiple myeloma were missing. The obvious similarities between the patients with autoimmune diseases and the patients with malignant monoclonal gammopathies suggest analogous pathogenetic mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Abortion, Spontaneous; Autoantibodies; Autoimmune Diseases; Blood Coagulation Factors; Blood Coagulation Tests; Brain Diseases; Female; Humans; Immunoglobulins; Lupus Coagulation Inhibitor; Lupus Erythematosus, Systemic; Lymphoma; Multiple Myeloma; Paraproteinemias; Pregnancy; Thromboplastin; Thrombosis

This study describes the use of a modified dilute thromboplastin assay for demonstration of the lupus coagulation inhibitor in plasma. A one-stage clotting inhibition assay is used in which the ability of the test plasma to prolong the clotting time of normal plasma is measured. The method is easy to perform and suitable for routine use. Clinical and laboratory data from five patients with the inhibitor but without systemic lupus erythematosus are presented. Thromboembolic manifestations were observed in three patients and obstetric complications possibly due to placenta thrombosis in two. One patient showed a bleeding tendency, associated with prothrombin deficiency. A number of pathological coagulation analyses and other laboratory data may be due to affinity of the lupus inhibitor for negatively charged phospholipids in vitro.

Topics: Abortion, Spontaneous; Adult; Anticoagulants; Blood Coagulation Factors; Blood Coagulation Tests; Female; Fetal Death; Humans; Lupus Coagulation Inhibitor; Pregnancy; Pregnancy Complications, Cardiovascular; Thromboembolism; Thromboplastin

Topics: Abortion, Spontaneous; Blood Coagulation; Blood Coagulation Tests; Catheterization; Curettage; Female; Fibrinolysis; Humans; Methods; Pregnancy; Thromboplastin; Time Factors; Uterine Diseases; Uterus

Topics: Abortion, Spontaneous; Adult; Afibrinogenemia; Blood Coagulation Tests; Female; Fetal Death; Fibrinogen; Heparin; Humans; Pregnancy; Thromboplastin

Topics: Abortion, Spontaneous; Adult; Antifibrinolytic Agents; Blood Coagulation Disorders; Decidua; Extraembryonic Membranes; Female; Fibrinolytic Agents; Humans; Muscle, Smooth; Phosphotransferases; Pregnancy; Thromboplastin; Uterus

Topics: Abortion, Spontaneous; Blood Coagulation Factors; Blood Coagulation Tests; Female; Hemostasis; Heparin; Humans; Pregnancy; Prothrombin Time; Thromboplastin