During a woman’s reproductive years, ovary-derived circulating estradiol (E2) and progesterone (P4) target the functional endometrial layer to induce cyclical changes that prepare it for implantation. Non-fertile cycles lead to sloughing of the endometrium in the menstrual fluid; restoration begins in association with hypoxia induced angiogenesis in the follicular phase of the next cycle as rising circulating E2 levels initiate cell proliferation. Following ovulation, an increase in circulating P4 levels stimulates E2-primed endometrial cells in the presence of a second peak of E2 to stop their proliferation and initiate differentiation. Consequently, endometrial glands become torturous and secrete an array of products that markedly affect the uterine milieu encountered by the implanting blastocyst while the luminal epithelium is prepared for blastocyst attachment. The stromal cells undergo decidualization, which represents a complex series of morphological and biochemical changes that transform precursor stromal cells into decidual cells. The decidualization reaction is initiated around blood vessels and under the glands [1].

Under continued E2 and P4 stimulation, decidualization spreads throughout the luteal phase endometrium [2]. Integral to the decidualization process is a marked alteration in the expression of proteins that promote hemostatsis and enhance vascular stability. Chief among these is tissue factor (TF), a 47 kDa trans-cell membrane glycolprotein member of the class-2 cytokine receptor family comprised of a hydrophilic extracellular domain, membrane-spanning hydrophobic domain and cytoplasmic tail. Under physiological conditions, endothelial cells do not express TF, whereas it is expressed constitutively at perivascular sites such as the adventitia of arteries and veins and in the parenchyma of various organs. Here TF forms a “hemostatic envelope” that protects against vascular injury by promoting binding of plasma-derived factor VII or its active form FVIIa to trans-cell membrane TF to trigger coagulation [3]. The clotting cascade depicted in Figure 1 indicates that TF/VIIa complex activates either FX (extrinsic pathway) or FIX (intrinsic pathway) to initiate a series of interactions among various clotting factors that culminate in proteolytic cleavage of prothrombin to thrombin, that then promotes hemostasis by forming the fibrin clot [4]. Figure 1 also indicates central roles for tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor type-one (PAI-1) in modulating hemostasis. Specifically, tPA acts as the primary fibrinolytic agent, which counteracts the prothrombotic

action of TF by binding to fibrin and cleaving Arg560- Val561 in its preferred substrate, plasminogen. The resulting generation of plasmin efficiently degrades fibrin. Its role as the fast inactivator of tPA enables PAI-1 to plays a critical role in determining the hemostatic balance [5].

The relative magnitude of TF expression conforms to a tissue-specific pattern. Specifically, low TF levels are present in the liver, spleen, skeletal muscle and thymus where hemostasis appears to depend primarily on the slower acting intrinsic coagulation pathway [6] (Figure 1). By contrast, the presence of high TF levels in the placenta, skin, heart, lung, brain and uterus is consistent with a requirement for accelerated hemostatic protection utilizing the faster extrinsic coagulation pathway. That no human genetic TF deficiency has been described indicates both the critical importance of TF expression to human survival while precluding direct evaluation of its absence in humans [6]. In mice, knocking out TF gene expression results in embryonic lethality. Specifically, TF null mice develop fragile vessels and die of hemorrhage in utero. However, incorporation of a human gene that expresses TF at only 1% of wild type levels rescues the knockout mice and produces live born pups. Adult low-TF mice exhibit hemostatic defects in tissues that normally express high levels of TF such as heart, lung, uterus and placenta [6]. In the mouse uterine wall, expression of high levels of TF in epithelial cells protects against hemorrhage during gestation as well as during the post-partum period. Pregnant low TF mice experience formation of blood pools in the placental labyrinth as well as high fatality rates postpartum [6].

Stimulation by circulating E2 and P4 continues to induce the decidualization reaction throughout human pregnancy. The crucial role played by the extrinsic coagulation pathway mediated by TF-expressing decidual cells in maintaining hemostasis during human pregnancy is evident in the events after implantation and subsequent placentation. Implantation of the blastocyst is followed by cytotrophoblast invasion of the decidua. Initially, cytotrophoblast breach endometrial capillaries and venules embedded in a matrix of decidual cells expressing high levels of TF. This invasive process institutes the primordial uteroplacental circulation and provides the embryo with a vital source of oxygen and nutrients while removing carbon dioxide and other waste products [7]. However, it also risks potential pregnancy-terminating hemorrhage as indicated by the phenomenon of “chemical pregnancy”, which is characterized by the transient appearance of trophoblast-derived HCG in maternal blood followed by local decidual bleeding [8,9].

Subsequent to these early pregnancy events, extravillous interstitial cytotrophoblast traverse the decidua to breach and remodel spiral arteries and arterioles into low-resistance, high-capacity vessels [10,11]. The resulting increase in uteroplacental blood flow provides vital nutrients and oxygen to the developing fetal-placental unit. Depending on the gestational age of occurrence, decidual hemorrhage during this period is associated with spontaneous abortion, abruption with or without stillbirth or preterm birth [12-14]. However, decidual cell-expressed TF is temporally and spatially positioned to promote local hemostasis thereby counteracting the threat of bleeding during both the initial and subsequent periods of endovascular cytotrophoblast invasion. Among cells at the maternal-fetal interface in human term decidua, immunoreactive TF levels are highest in decidual cells [15]. By contrast, adjacent interstitial trophoblasts are essentially devoid of TF immunostaining [15]. Thus, continued elevation of TF expression by decidual cells protects against the hemostatic demands of delivery both preterm and at term.

In addition to promoting hemostasis, the generation of thrombin by TF also affects a multitude of biological functions through thrombin’s binding to and activation of members of the protease activated receptor (PAR) family, which is comprised of four distinct “seven membrane” G protein coupled receptors. Thrombin promotes proliferation and chemotaxis of immune and mesenchymal cells and activates endothelial cells via PAR-1, 3 and 4, whereas PAR-2 acts as a receptor for trypsin and trypsinlike enzymes including the TF/VIIa complex. In addition, PAR-1 and PAR-2 can be activated by FXa [16]. As exemplified by thrombin, after each PAR ligand binds to its specific receptor, its serine protease domain cleaves that PAR to expose an N-terminal tethered ligand domain that autoactivates the cleaved receptor [17]. Previously, our laboratory found that stromal cells from cycling human endometrium express PAR-1 [18] and that incubation of such cultured stromal cells with thrombin or TRAP, a synthetic peptide activator of PAR-1, enhances secreted levels of the primary angiogenic agent vascular endothelial growth factor [19]. Moreover, as detailed below, cultured third trimester decidual cells continue to respond to thrombin with enhanced expression of PAR-mediated endpoints implicated in the genesis of preterm premature rupture of the membranes (PPROM) and preterm delivery (PTD).

Taken together, these observations indicate that excess thrombin derived from decidual cell expressed TF acts as an autocrine/paracrine effector that induces abruptionassociated PTD by inducing functional P4 withdrawal in decidual cells. These findings complement our earlier observations indicating that such decidual cell-derived thrombin also elicits PPROM and PTD by acting in autocrine/paracrine fashion to enhance expression of decidual ECM degrading proteases [41,42], by inducing expression of the neutrophil chemoattractant and activator, IL-8 [43] and by directly weakening the amnion [49].