Platelet’s functions and their role in the coagulation process in pregnancy are well documented, and there are several data to suggest their importance in embryo implantation, placental development, preeclampsia, HELLP syndrome, and other physiological and pathophysiological processes.
Platelets may play a role in placentation
Circulating platelets stores several bioactive mediators within their granulas, including PDGF-B, bFGF, HGF, IGF, IGF-1, PF4, TSP1, TGF-β1 and VEGF, and they release them upon stimulation. Most of these molecules are found to be relevant in mouse placentation, for instance bFGF, IGF-1 or PDGF which were proved to be an effective protector of trophoblast cells against TNF-α and INF-γ induced apoptosis
in vitro[
48]. Beyond that, the deficit of PDGF-B or PDGF-R-β, results vessel dilation in the placenta[
49]. Moreover the developmental failure of the placenta is associated with the absence of lipid-phosphatase 3 (LLP-3)[
50]. Such findings implemented the necessity of
in vivo animal experiments to further investigate the role of platelets in implantation and development. The mouse model of quantitative platelet disorder (NF-E2 deficient mice) and qualitative thrombocyte defect (Gαq, Par-3, or Par-4 deficient mice) makes them eligible for such task.
The NF-E2 transcriptional factor is a crucial protein of the megakaryopoiesis, therefore NF-E2 “knock-out” mice develops a severe quantitative platelet deficiency[
51]. Such condition in these animals is associated with significant intrauterine growth retardation (IUGR), explained by the vascular abnormality of the placental labyrinthine layer, due to impaired blood vessel maturity. Interestingly, failure of the placental vascularization is only reported in NF-E2 null embryos, but not in NF-E2 null mothers, nor mice with other platelet disorders. NF-E2 “knock-out” mothers delivers same number of litter, and the embryonic loss was found to be not significantly different compared to wild type mice, despite that histological evaluation revealed numerous spacious blood pools in the placenta (intraplacental hemorrhage)[
52,
53].
On the other hand, in case of qualitative thrombocyte abnormalities placental disorders can not be revealed. For example Gαq-deficient mice (lack of the α-subunit of the guanine nucleotide binding protein Gq), has normal number of thrombocytes, but their platelets can not be activated, develop normal placental development and embryos[
54]. Likewise, when protease-activated receptor 3 and 4 (Par-3 and Par-4) mediated platelet activation is also missing in mice an intact, normal placenta and embryo develops[
55].
Next to the experimental results from mice, human data also show that maternal platelets might play important role in the placental vascular remodeling, which assures proper placental perfusion and maintains the pregnancy itself. Such phenomenon is based on the endovascular invasion of trophoblast cells into the maternal spiral arteries, enabling them to transform large-caliber vessels[
56]. The mechanism of such alteration is, that maternal platelets are trapped in the lumen of the spiral arteries by endovascular trophoblast aggregates. These activated platelets are communicating with neutrophil granulocytes, by expressing P-selectin on their surfaces. These cell surface cell adhesion molecules are binding to their ligands (P-selectin glycoprotein ligand-1; PSGL-1) on the activated granucolcytes, and in turn they produce PAF to facilitate the further aggregation processes[
57]. Extravillous trophoblast cells express chemokine receptor (CCR1), which plays as a functional receptor for trophoblasts. Their ligands (CCR1 ligand) are secreted by the activated platelets in the placenta to encourage the remodeling processes, and to attract more endovascular trophoblasts to the site[
58]. To achieve such vascular caliber changes during the early placentation a direct contact between the platelets and the trophoblast cells is not fully required. Therefore it is suspected that some soluble platelet-derived factor(s) are regulating the placental vascular remodeling processes in human[
58].
Clinical observations, in case of recurrent pregnancy loss, intrauterine growth restriction and preeclampsia, highlighted the effectiveness of low-dose platelet cyclooxygenase inhibitor (Aspirin®), which reduces the platelet aggregation by suppressing the TXA2 synthesis[
59]. However, pregnant women with Bernard-Soulier syndrome or other congenital thrombocytopenias usually have an uncomplicated pregnancy and they deliver a healthy newborn, but the risk for a potential postpartum bleeding is increased[
60].
Platelets in preeclampsia and HELLP syndrome
Pregnancy is a condition of immunological balance between the mother and her semi-allograft fetus. Besides, pregnancy is considered as a controlled pro-coagulation, and a mild inflammatory state. As platelets are involved in all of these processes, their consumption is enhanced. Consequently, platelet production is augmented, resulting in a slightly increased ratio of young platelets in the bloodstream. However, platelet count remains normal (150,000 to 450,000 platelets per microliter) throughout normal pregnancy. This slim equilibrium will be disturbed in conditions where complex maternal adaptation to her pregnancy is inadequate.
Preeclampsia is a serious disease in pregnancy with heterogeneous etiology[
61,
62]. Early-onset, or hypovolemic, or serious preeclampsia and the hemolysis, elevated liver enzymes, and low platelet count (HELLP) syndrome are thought to be conditions due to imbalance between the mother and fetus[
63]. Theoretically, the progenitors could be of normal fetal antigens but their pathologically increased mass in the maternal circulation and/or the abnormal maternal reaction to them (“
maladaptation”) could lead to the clinical symptoms. Platelets play a pivotal role in the holistic defending mechanism, being stimulated in increasing degrees in these serious diseases. Overstimulation collapses this platelet-associated regulation contributing to the development of end-organ damages.
Early-onset preeclampsia can be considered as a 2-stage disease. Abnormal placentation (first stage) through endothelial damage is responsible for the potential of end-organ manifestations (second stage). The first stage is characterized by effects of anti-angiogenic substances such as the soluble fms-like tyrosine kinas-1 (sFlt-1)[
64], endoglin (sEng)[
65], and human interferon-inducible protein 10 (IP-10 or CXC10)[
66] which bind and neutralize different growth factors, placental growth factor (PlGF), VEGF, and TGF-β required for placental and fetal angiogenesis. Agents directly from this shallowly implanted placenta and also from activated leukocytes or platelets may represent the link between abnormal placentation and endothelial inflammatory injury. Recent candidates are the anti-angiogenic agents; free oxygen radicals and some cytokines, first of all TNF-α[
67,
68], thrombogenic content of miroparticles released from the surface of different cells[
69]; syncytiotrophoblast microvilli or vesicles[
70] releasing sFlt-1 and sEng; fetal cells and cell-free fetal DNA[
71] circulating in a relatively huge amount in preeclamptic maternal bloodstream. Consequently, markers of endothelial injury e. g. fibronectin[
72] soluble thrombomodulin[
73], or von Willebrand factor[
74] show elevated levels in preeclampsia. Similarly, levels of platelet-aggregation products (e.g. β thromboglobulin, PF4, TXA2) are elevated[
75].
Epidemiological studies suggested that “immune-maladaptation” may have an etiologic role in the development of preeclampsia[
76]. Long (>4 months) maternal ejaculate-exposition results in slow alloimmunization, and development of immuntolerance against the given gentile antibodies[
71]. In the absence of appropriate partner-specific mucosal immuntolerance the incidence and severity of preeclampsia will increase[
77]. Fetal proteins contact with maternal immune system not only at the feto-maternal interface but also in the maternal circulation, where fetal cells, cell debris, or DNA can be detected in huge number in preeclamptic individuals, showing correlation with the severity of the disease[
71]. Platelets play a pivotal role in the initiation and modulation of immunological processes. Soluble platelet-released immune modulators (chemokines, cytokines) coordinate leukocyte migration, endothelial adhesion, and function[
78]. There is no doubt that signals of activated platelets are essentials in the development of maternal immune-respose in preeclampsia too.
It is well known for some decades that platelets and also intravascular coagulation cascade are activated in preeclampsia. Damaged endothelial cells produce more adhesive substances (e.g. fibronectin, vascular cell adhesion molecule-1, E-selectin) and possess less antithrombotic capacity (e.g. weak thrombomodulin effects) than normal endothelial cells[
73,
79]. Impairment of endothelium exposes the subendothelial collagen, a potent platelet-activating agent, to vessel content. Increased nitric oxide (NO) production during normal pregnancy effectively decreases the sensitivity of platelets to pro-aggregating agents[
80]. Asymmetric dimethyl arginine (ADMA), the endogenous inhibitor of NO synthesis, levels are increased in preeclampsia[
81] lowering the effectiveness of an important protective mechanism of platelet activation. In accordance, the augmentation of endogenous and reactive NO production shows beneficial effects in gestational hypertension[
82]. Similarly, damaged endothelial cells produce less prostaglandin I
2 (PGI
2 or prostacyclin), another potent agent against platelet activation[
83].
Activated platelets not only release the vasoconstrictor TXA
2, but also increase the expression of adhesive agents[
84]. Activated platelets this way enhance further platelet adhesion, and also the endothelium − leukocyte contact and trigger leukocyte arrest and transendothelial migration[
85].
Erythrocyte deformability has been found to be decreased in preeclampsia which may contribute to decreased capillary circulation[
86], since capillary diameter is about the half of those of red blood cells. Peripheral mechanical hemolysis is a hallmark of HELLP but characteristic feature of severe preeclampsia as well[
87]. Hemolysis in preeclampsia may further enhance the platelet activation since breaking red blood cells release ADP[
88]. Besides, damaged erythrocytes show an enhanced aggregability as well[
89]. This way altered hemostasis with activated platelets markedly contributes to the collapse of microcirculation resulting in tissue hypoperfusion with subsequent end-organ dysfunction in serious preeclampsia.
Platelets actively take part in the defensive action against pathogenic microorganisms. Platelets quickly accumulate and become activated at the site of invasion[
90]. They not only release effective agents to damage invading cells but also capable of eliminate them from the circulation. The cationic thrombocidins, releasing from the α granules, are able to depolarize bacterial membranes and inhibit the synthesis of some intracellular molecules[
91]. Besides, secondary to stimuli by ADP or stromal cell-derived factor-1 platelets can eliminates bacteria or viruses by internalization (“engulfment”)[
92]. The relationship between preeclampsia and infection was posed by the preeclampsia model based on low-dose endotoxin infusion in pregnant rats[
93]. Since then several studies found correlation between preeclampsia/HELLP syndrome and different forms of infection:
-
Concomitant urogenital infection during pregnancy enhances the incidence of preeclampsia[
94];
-
Statistical data confirm that preeclampsia incidence increases when patients had
Chlamydia pneumoniae infection previously[
95];
-
Periodontitis during pregnancy predisposes for preeclampsia[
96];
-
Use of antibiotics during pregnancy decreases the incidence of preeclampsia[
97];
-
In patients with HELLP syndrome a concomitant infection is frequent[
98];
It is tempting to assume that the link between preeclampsia and different inflammations may be further enhancing of platelet activation. This presumption seems to be supported by the modest but obvious efficacy of aspirin in the prevention of preeclampsia[
99]. Besides, bacterial endotoxin in known to destroy endothelial cells enhancing their dysfunction. Furthermore, platelet adhesion and aggregation could be the consequence of elevated shear stress[
100]. High shear stress is ordinarily caused by hypertension, which is a criterion of preeclampsia.
Platelet activation in HELLP syndrome has similar background than it is characteristic for severe preeclampsia; however, there are some differences. The early trophoblast lesion is more pronounced in HELLP. The antiangiogenic sEng levels are significantly higher in HELLP than in preeclampsia. Development of microangiopathy, secondary to increased sEng, sFlt-1, activated TNFα, and von Willebrand factor in HELLP may explain the extreme platelet consumption as consequence of overstimulation[
63].
In severe preeclampsia and HELLP syndrome platelets are highly stimulated. Falling the platelet count and elevation of platelet aggregation products precede the manifestation of clinical symptoms by several weeks[
75]. This stimulation could be enhanced further by concomitant infection which worsens the condition. Inhibition of platelet aggregation, starting in first trimester, may have a beneficial effect. On the contrary, when the consumption permanently exceeds the production the platelet-associated regulatory system (PARS) collapses and the condition turns to critical state.