Third-trimester Placentas Research Articles

Effect of THP-1 Cells on the Formation of Vascular Tubes by Endothelial EA.hy926 Cells in the Presence of Placenta Secretory Products.

We studied the effect of THP-1 cells on the formation of vessel-like structures by endothelial cells in the presence of placenta-conditioned media. Addition of THP-1 cells to endothelial cells cultured in the presence of media conditioned by first-trimester placentas led to an increase in the length of cell tubes and reduced their number in comparison with endothelial cell monoculture. In the presence of media conditioned by third-trimester placentas, THP-1 cells did not affect the length and number of cell tubes formed by endothelial cells. When evaluating the formation of vessel-like structures by endothelial cells in co-culture, marked decrease in the length of cell tubes in the presence of media conditioned by first-trimester placentas vs. third-trimester placentas was noted. No differences in the length and number of cell tubes formed by endothelial cells co-cultured with THP-1 cells in the presence of placental factors from women with preeclampsia and uncomplicated pregnancy were found. These findings can reflect the peculiarities of the influence of macrophages on the formation of blood vessels by endothelial cells in the placenta.

Telomere homeostasis in placentas from pregnancies with uncontrolled diabetes

ObjectiveDiabetes during pregnancy causes an intrauterine environment that influences lifetime sickness of the mother and the fetus. There is a correlation between diabetes and telomere shortening; however, very little is known about telomere homeostasis in the placenta. We aimed to study the telomerase complex in placentas and in cord blood leukocytes from patients with poorly controlled diabetes. MethodsBiopsies from 16 third-trimester placentas and cord blood samples from pregnancies complicated with uncontrolled diabetes and from 16 gestational age-matched controls from uncomplicated pregnancies were examined. The expression of hTERT (human telomerase reverse transcriptase) was evaluated by immunohistochemistry and by RT-RCR. TERC gene copy number and telomere capture were evaluated by FISH. ResultsTelomerase expression was significantly lower in the diabetic placentas, both the protein (17.8 ± 2.8% cellular staining vs. 37 ± 5.32%, P = 0.012) and the mRNA levels (0.42 ± 0.03 folds, P = 0.022). Lower expression of TERC gene copy number were shown in the diabetic placentas compared to the healthy controls (1.7 ± 0.8% vs. 3.7 ± 1.6%, P = 0.035). We also detected higher percentage of cells with telomere capture among the diabetic trophoblasts compared to the healthy controls (19.8 ± 5.12% vs. 9.6 ± 3.65%, P = 0.038). Those differences were not observed in cord blood leukocytes from the same samples. ConclusionsUncontrolled diabetes during pregnancy disrupts telomere-telomerase homeostasis in the trophoblasts. These changes may increase the risk for metabolic diseases in adulthood among offspring of pregnancies complicated by gestational diabetes mellitus as part of intrauterine programming. These variations were not observed in cord blood leukocytes, which imply different telomere homeostasis mechanisms in fetal cord blood.

Genomic Alterations Are Enhanced in Placentas from Pregnancies with Fetal Growth Restriction and Preeclampsia: Preliminary Results

Fetal growth restriction (FGR) secondary to placental insufficiency and preeclampsia (PE) are associated with substantially increased childhood and adult morbidity and mortality. The long-term outcomes are related to placental aberrations and intrauterine programming. Advances in microarray technology allow high-resolution, genome-wide evaluation for DNA copy number variations - deletions and duplications. The aim of our study was to demonstrate the usefulness of microarray testing in FGR placentas. Using Affymetrix GeneChip for chromosomal microarray (CMA), we analyzed 10 placentas from pregnancies with FGR attributed to placental insufficiency; 5 with FGR below the 5th percentile and 5 from the 5th to <10th percentiles. All fetuses had normal anomaly scans and karyotypes. We also analyzed 5 third-trimester placentas from pregnancies complicated by PE with severe features and 5 from PE without severe features, all with appropriately grown fetuses. The results were compared to 10 placentas from uncomplicated pregnancies with healthy neonates. CMA analysis identified more genomic alterations in FGR (p < 0.05) and in PE (p < 0.05) placentas than in healthy controls. There was a correlation to the severity of FGR and PE. The genomic alterations were below the resolution of normal karyotyping. The altered genes are related to adult human height, stress reactions and to cellular migration, differentiation and adhesion. Though very preliminary, our data support evaluating FGR and PE placentas using CMA. Larger data sets are needed for further evaluation of the findings and their clinical implications.

Immunizing with mouse trophoblast to obtain mouse-human cross-reacting antibodies against normal and neoplastic human trophoblast.

Monoclonal antibodies raised against living, early invasive mouse trophoblast cells were screened on paraffin sections from first- and third-trimester placentas and from hydatidiform moles and choriocarcinoma. Several mouse-human cross-reacting antibodies were recognized, which implies that mouse trophoblast cells can be used as immunogen for producing antibodies against human trophoblast. Among the new antibodies obtained, some were selected for further study. That panel includes a trophoblast specific antibody with capacity to differ between invasive and noninvasive molar tissues, and two antibodies, which detect antigen epitopes in the normal, but not in the neoplastic trophoblast.

Trophoblast expression dynamics of the tumor suppressor gene gastrokine 2.

Gastrokines (GKNs) were originally described as stomach-specific tumor suppressor genes. Recently, we identified GKN1 in extravillous trophoblasts (EVT) of human placenta. GKN1 treatment reduced the migration of the trophoblast cell line JEG-3. GKN2 is known to inhibit the proliferation, migration and invasion of gastric cancer cells and may interact with GKN1. Recently, GKN2 was detected in the placental yolk sac of mice. We therefore aimed to further characterize placental GKN2 expression. By immunohistochemistry, healthy first-trimester placenta showed ubiquitous staining for GKN2 at its early gestational stage. At later gestational stages, a more differentiated expression pattern in EVT and villous cytotrophoblasts became evident. In healthy third-trimester placenta, only EVT retained strong GKN2 immunoreactivity. In contrast, HELLP placentas showed a tendency of increased levels of GKN2 expression with a more prominent GKN2 staining in their syncytiotrophoblast. Choriocarcinoma cell lines did not express GKN2. Besides its trophoblastic expression, we found human GKN2 in fibrotic villi, in amniotic membrane and umbilical cord. GKN2 co-localized with smooth muscle actin in villous myofibroblasts and with HLA-G and GKN1 in EVT. In the rodent placenta, GKN2 was specifically located in the spongiotrophoblast layer. Thus, the gestational age-dependent and compartment-specific expression pattern of GKN2 points to a role for placental development. The syncytial expression of GKN2 in HELLP placentas might represent a reduced state of functional differentiation of the syncytiotrophoblast. Moreover, the specific GKN2 expression in the rodent spongiotrophoblast layer (equivalent to human EVT) might suggest an important role in EVT physiology.

What is the role of matrix metalloproteinase-2 in placenta percreta?

This study compared the placental expression of the matrix metalloproteinase-2 (MMP-2) enzyme, which is thought to play a key role in the penetration of trophoblastic cells, in third-trimester placenta percreta (PP) patients with that of women with normal pregnancies. Twenty-five pregnant subjects who underwent cesarean section due to PP and 25 term pregnant subjects who underwent cesarean section for obstetric reasons were included in the study. Demographic data, pathology reports, and histopathological samples were examined. Blocks containing samples of placenta underwent immunohistochemical analysis using the MMP-2 antibody. Immunohistochemical expression of placental samples obtained from both groups was examined and compared. There were no statistically significant differences in the demographic data (P > 0.05). With regard to immunohistochemistry, cytoplasmic staining of trophoblastic cells was considered immunohistochemically positive. In the PP tissue samples, positive MMP-2 staining was detected as follows: 0 immunoreactivity, one patient (4%); 1(+), six patients (24%); 2(+), seven patients (28%); and 3(+), 11 patients (44%). In the term pregnant placental tissue samples, positive MMP-2 staining was detected in five patients (20%) at 0 immunoreactivity, 12 (48%) at 1(+) immunoreactivity, five (20%) at 2(+) immunoreactivity, and three patients (12%) at 3(+) immunoreactivity. Immunohistochemical expression was significantly different between the PP and normal term pregnancy placental tissues (P = 0.02). The stronger expression of the MMP-2 enzyme in the PP as compared to the normal placental tissue suggests that this enzyme may be an effective mediator in the pathogenesis of PP.

Myostatin is localized in extravillous trophoblast and up-regulates migration.

Myostatin is a highly conserved secretory protein that negatively regulates muscle development by affecting both proliferation and differentiation of muscle cells. In human placentae the expression of myostatin is negatively correlated with gestational age, and in placental explants, myostatin acts to facilitate glucose uptake. Myostatin expression is known to be higher in the placentae of pregnancies complicated by preeclampsia. Proper placental development is crucial for a healthy and successful pregnancy. Alterations to the function of the placental cells after treatment with myostatin have not previously been published. This study investigated the localization of myostatin in extravillous trophoblast (EVT) of human placentae. Furthermore, the effect of myostatin treatment on the proliferative and migrative capabilities of these placental cells was investigated. Myostatin is localized in EVT, as identified by the immunohistochemistry of third-trimester placentae and immunocytochemistry of first-trimester EVT isolations positively staining for myostatin and human leukocyte antigen-G. Treatment of an EVT cell line (HTR-8/SVneo) and primary isolated EVT with varied concentrations of myostatin resulted in a significant increase in the proliferation (HTR-8/SVneo; P < .0001) and migration (HTR-8/SVneo and primary isolated EVT; P < .05), with proliferation being dose dependent and migration being dose independent. Myostatin localization was positively identified in EVT. Myostatin positively affected proliferation (HTR-8/SVneo) and migration of EVT (HTR-8/SVneo and primary isolated EVT). For the first time, the effect of myostatin treatment on placental cells is described. The results provide a base from which further in vitro investigations on myostatin's ability to modulate placental cell function can be made.

Epigenome-wide DNA methylation assay reveals placental epigenetic markers for noninvasive fetal single-nucleotide polymorphism genotyping in maternal plasma.

The use of DNA methylation difference between maternal blood cell and fetal (placental) DNA is one of the main areas of interest for the development of fetal epigenetics markers in maternal plasma. We employed a methylation array (HumanMethylation450 array, Illumina, Inc.) to identify novel biomarkers that are specially hypermethylated in placental DNA versus maternal blood cells in a genome-wide basis. Validation by bisulfite genomic sequencing was performed and the priority was given to potential targets that harbor differential methylated CpG sites overlapped with at least two methylation-sensitive restriction enzyme (MSRE) recognizing sites, as well as one polymorphic single-nucleotide polymorphism (SNP), within a short DNA stretch. Three candidate regions of PSMB8, SKI, and CHST11 gene were selected for developing a preliminary polymerase chain reaction assay with MSRE digestion of maternal plasma DNA. SNP genotypes were confirmed by direct sequencing. We identified 2944 and 5218 fetal-specific hypermethylated CpG sites in the first- and third-trimester placenta, respectively, of which 2613 were overlapped, suggesting a consistency of differential methylation during the whole pregnancy. The array results were confirmed by bisulfite genomic sequencing. The preliminary tests in maternal plasma showed that postdigestion hypermathylated versions of these candidate molecules were detectable only in pregnant women. We further revealed that methylated targets in maternal plasma possessed the fetal SNP genotypes. The present studies systematically identified hypermethylated sites in fetal tissues and preliminarily demonstrated that some of the fetal epigenetic markers that contain informative SNPs have great potential for noninvasive fetal genetic diagnosis.

Decreased expression and DNA methylation levels of GATAD1 in preeclamptic placentas

Expression of syncytin-1, or the human endogenous retroviral family W member 1 (HERVWE1) in human placental trophoblasts is regulated by DNA methylation. Increased DNA methylation and decreased expression of syncytin-1 have been observed in preeclamptic placentas. The syncytin-1-mediated fusogenic as well as non-fusogenic activities, e.g., cell cycle promotion, anti-apoptosis, and immune suppression, are implicated in the pathogenic changes in preeclamptic placentas. It is noteworthy that in a close vicinity to syncytin-1 there are two genes, peroxisome biogenesis factor 1 (PEX1) and GATA zinc finger domain containing 1 (GATAD1), as well as multiple CpG islands around these genes. In this study we determined if these adjacent genes might, like syncytin-1, subject to epigenetic regulation in preeclamptic placentas. Data from quantitative real-time PCR and Western blotting indicated that while PEX1 expression remained stable, GATAD1 expression was significantly decreased in the third-trimester placentas associated with preeclampsia than those associated with normal pregnancy. Immunohistochemistry detected high GATAD1 expression in trophoblast linage, and confirmed its reduced levels in preeclamptic placentas. However, COBRA and bisulfate sequencing detected decreased DNA methylation in levels in the 3 [prime] region of GATAD1 gene in preeclamptic placentas. The positive correlation between 3 [prime] methylation and GATAD1 expression was confirmed by treatment of choriocarcinoma JAR cells with DNMT inhibitor. These data pointed to a potential role of GATAD1 for the syncytium deficiency often associated with preeclamptic placentas. The sharp contrast of the methylation alterations for the closely positioned GATAD1 and HERVWE1 may provide a useful model for studying the accurate control of DNA methylation as well as their positive and negative impact on gene expression in placental trophoblasts.

Placental hydroxymethylation vsmethylation at the imprinting control region 2 on chromosome 11p15.5

In addition to methylated cytosines (5-mCs), hydroxymethylcytosines (5-hmCs) are present in CpG dinucleotide-enriched regions and some transcription regulator binding sites. Unlike methylation, hydroxymethylation does not result in silencing of gene expression, and the most commonly used methods to study methylation, such as techniques based on restriction enzymatic digestion and/or bisulfite modification, are unable to distinguish between them. Genomic imprinting is a process of gene regulation where only one member of an allelic pair is expressed depending on the parental origin. Chromosome 11p15.5 has an imprinting control region (ICR2) that includes a differentially methylated region (KvDMR1) that guarantees parent-specific gene expression. The objective of the present study was to determine the presence of 5-hmC at the KvDMR1 in human placentas. We analyzed 16 third-trimester normal human placentas (chorionic villi). We compared two different methods based on real-time PCR after enzymatic digestion. The first method distinguished methylation from hydroxymethylation, while the other method did not. Unlike other methylation studies, subtle variations of methylation in ICRs could represent a drastic deregulation of the expression of imprinted genes, leading to important phenotypic consequences, and the presence of hydroxymethylation could interfere with the results of many studies. We observed agreement between the results of both methods, indicating the absence of hydroxymethylation at the KvDMR1 in third-trimester placentas. To the best of our knowledge, this is the first study describing the investigation of hydroxymethylation in human placenta using a genomic imprinting model.

Imbalance of expression of bFGF and PK1 is associated with defective maturation and antenatal placental insufficiency

ObjectiveDefective placental maturation is associated with restricted functional capacity and adverse perinatal fetal outcomes. The aim of the study was a comparative analysis of the role of mRNA expression of various angiogenic factors in placental maturation defects. Study designWe examined the mRNA expression patterns of prokineticin 1 (PK1), its receptors (PKRs), basic-fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) in tissue from third-trimester placentae that exhibited delayed or accelerated villous maturation. ResultsThe expression of PK1 and PKR2 was elevated in placental tissue exhibiting accelerated maturation and a predominant differentiation of terminal villi. The opposite was found in tissue exhibiting delayed maturation and deficiency of the terminal villi. In addition, low expression of bFGF correlated with the predominant differentiation of terminal villi, whereas the opposite was observed when terminal villi were deficient. The expression of VEGF, PIGF, and PKR1 showed no significant differences between the groups. ConclusionDefective placental maturation is associated with an imbalance of expression of bFGF and PK1. Our results demonstrate an involvement of the PK1/PKR2-signalling pathway in the regulation of the functional adequate capillarization in late pregnancy. We propose the bFGF/PK1-ratio as a monitor of placental function and a possible indicator of latent clinical problems, such as placental dysfunction leading to fetal hypoxia.

The chemokine CXCL6 restricts human trophoblast cell migration and invasion by suppressing MMP-2 activity in the first trimester

Can the chemokine CXCL6 affect trophoblast cell migration and invasion in human first-trimester placenta? Chemokine CXCL6 inhibits trophoblast cell migration and invasion by suppressing matrix metalloproteinase (MMP)-2 activity in human first-trimester placenta. Several chemokines including CXCL8, CXCL12, CXCL14, CXCL16, CX3CL1, CCL14 and CCL4 can promote or inhibit trophoblast cell migration and invasion in human first-trimester placenta. We used the trophoblast cell line HTR8/SVneo cells, primary trophoblast cells and villi explants to investigate the effect of rhCXCL6 on trophoblast cell migration and invasion. First, the CXCL6 RNA transcript level was detected in HTR8/SVneo cells derived from human first-trimester, second-trimester and third-trimester placenta by RT-PCR. Protein expression of CXCL6 and its receptors was tested in first-trimester placenta by immunohistochemistry. Secreted CXCL6 protein was detected in HTR8/SVneo cell supernatants by enzyme-linked immunosorbent assay. Secondly, the effect of rhCXCL6 on HTR8/SVneo cell proliferation was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Thirdly, the effect of rhCXCL6 on cell migration and invasion of HTR8/SVneo cells, primary trophoblast cells and villi explants was tested by transwell migration and invasion assays, respectively. Last, MMP-2 and MMP-9 activity in the supernatants of HTR8/SVneo and primary trophoblast cells treated by rhCXCL6 in the invasion assay was assessed by gelatin zymography. Abundance of the CXCL6 RNA transcript increased with pregnancy development. CXCL6 and its receptor were expressed in several cells at the human maternal-fetal interface. RhCXCL6 inhibited trophoblast cell migration and invasion by suppressing MMP-2 activity. These experiments are only in vitro. According to the literature, CXCL6 could promote tumour cell migration and invasion by accelerating MMP-9 activity. However, CXCL6 inhibited trophoblast cell migration and invasion by suppressing MMP-2 activity in human first-trimester interface. These data suggest that strict regulation of CXCL6 is required for normal migration and invasion of cells, such as those involved at the maternal-fetal interface.

Cervical length and risk of antepartum hemorrhage in presence of low-lying placenta

Objectives: To evaluate whether transvaginal ultrasound cervical length (TVU CL) can predict antepartum bleeding (APB) in women with low-lying placenta. Study design: A retrospective study was performed including pregnancies with low-lying placenta for which third trimester TVU CL was available. Multiple pregnancies were excluded. Short cervix was defined as TVU CL ≤25 mm. Outcomes of interest were compared with respect to the TVU CL. Results: Forty three cases of singleton pregnancies complicated by low-lying placenta in third trimester were identified. Short cervix was reported in 8 cases (19%). APB (75% vs. 31 %, p = 0.02), blood transfusions (25% vs. 3%, p = 0.02), lower birth weight (2246 vs. 2985 g, p = 0.02), and neonatal intensive care unit (NICU) admissions (50% vs. 17%, p = 0.04) were more frequent in the women with short cervix. Rate of unplanned cesarean delivery for APB was similar between both the groups (25% vs. 28%, p = 0.83). Conclusions: In women with low-lying placenta persisting into third trimester, short cervical length can be used as a predictor for APB.

Expression and Localization of TLR4 and Its Negative Regulator Tollip in the Placenta of Early-Onset and Late-Onset Preeclampsia

Objective. Toll-like receptor 4 (TLR4) is a key component of the innate arm of the immune system that mediates inflammatory responses following exposure to bacterial lipopolysaccharides. In doing so, TLR4 may contribute to the pathogenesis of preeclampsia (PE). We sought to assess the spatio-temporal expression of TLR4 and its negative regulator Toll-interacting protein (Tollip) in placental tissues from normal and preeclamptic pregnancies. Methods. Using immunohistochemistry and immunoblotting, we investigated the localization of TLR4 and its negative regulator Tollip in first- and third-trimester human placenta. Results. TLR4 was found to be expressed in the cytoplasm of trophoblasts in both first- (n = 6) and third-trimester (n = 24) placental villous samples. Tollip was mainly located in the cytoplasm of cytotrophoblasts in the first-trimester placental tissues; in contrast, our analyses of third-trimester placental tissues demonstrated that Tollip was mainly located in the decidual cells. Using western blot analysis, TLR4 expression was shown to be significantly higher in early-onset PE tissues than control group placentas (n = 8 for each group, p < 0.01). However, we were unable to detect a significant difference between late-onset PE and normal controls (n = 8 for each group, p = 0.119). Conclusion. These data demonstrate the novel observation that TLR4 may play a much more important role in early-onset PE. Interestingly, the spatial expression of Tollip at different stages of gestation may play an important role in the pathophysiology of PE.

Effect of Macrophage Migration Inhibitory Factor (MIF) in Human Placental Explants Infected with Toxoplasma gondii Depends on Gestational Age

Because macrophage migration inhibitory factor (MIF) is a key cytokine in pregnancy and has a role in inflammatory response and pathogen defense, the objective of the present study was to investigate the effects of MIF in first- and third-trimester human placental explants infected with Toxoplasma gondii. Explants were treated with recombinant MIF, IL-12, interferon-γ, transforming growth factor-β1, or IL-10, followed by infection with T. gondii RH strain tachyzoites. Supernatants of cultured explants were assessed for MIF production. Explants were processed for morphologic analysis, immunohistochemistry, and real-time PCR analysis. Comparison of infected and stimulated explants versus noninfected control explants demonstrated a significant increase in MIF release in first-trimester but not third-trimester explants. Tissue parasitism was higher in third- than in first-trimester explants. Moreover, T. gondii DNA content was lower in first-trimester explants treated with MIF compared with untreated explants. However, in third-trimester explants, MIF stimulus decreased T. gondii DNA content only at the highest concentration of the cytokine. In addition, high expression of MIF receptor was observed in first-trimester placental explants, whereas MIF receptor expression was low in third-trimester explants. In conclusion, MIF was up-regulated and demonstrated to be important for control of T. gondii infection in first-trimester explants, whereas lack of MIF up-regulation in third-trimester placentas may be involved in higher susceptibility to infection at this gestational age.

Expression of laminin receptor 1 in human placentas from normal and preeclamptic pregnancies and its relationship with the severity of preeclampsia

To investigate the expression of laminin receptor 1 (LR1), a non-integrin-type laminin receptor, in preeclamptic and normal third-trimester placentas, as well as to investigate whether its expression differs with disease severity. Third trimester placental samples obtained from deliveries of preeclamptic (n=34) and normotensive healthy pregnant women (n=35) were immunohistochemically studied for the expression of LR1. The placentas from both mild (n=14) and severe (n=20) preeclamptic pregnancies were further assessed for strength of LR1 expression according to disease severity. When compared with normal placentas, the staining with LR1 protein in cytotrophoblasts and syncytiotrophoblasts was lower in preeclamptic placentas (P<0.05 and P<0.01, respectively). The intensity of staining with LR1 in decidual cells, cytotrophoblasts, syncytiotrophoblasts, and extracellular matrix cells of preeclamptic placentas did not vary with disease severity (P>0.05). Decreased LR1 expression in cytotrophoblasts and syncytiotrophoblasts of preeclamptic placentas, which may be independent of disease severity, might have a role in shallow trophoblastic invasion in preeclampsia.

Sonographic Maturation of the Placenta at 30 to 34 Weeks Is Not Associated With Second Trimester Markers of Placental Insufficiency in Low-risk Pregnancies

ObjectiveAdvanced placental maturation (Grannum [G] grade 3) before term is associated with adverse perinatal outcomes associated with placental insufficiency. The nature and timing of the underlying pathology of this process is presently unclear. We hypothesized that advanced placental maturation at 30 to 34 weeks’ gestation is not associated with established second trimester markers of severe placental dysfunction. MethodsIn a cohort study of 1238 low-risk Caucasian women with singleton pregnancies who had sonographic assessment of placental maturation and fetal growth at 34 weeks, the results of maternal serum screening (MSS) and uterine artery Doppler (UtAD) flow studies at 16 weeks were related to adverse perinatal outcomes associated with placental insufficiency: antepartum hemorrhage, preeclampsia, preterm birth < 37 weeks, small for gestational age (<10th percentile), or postnatal evidence of intrauterine growth restriction (IUGR; ponderal index < 5th percentile). ResultsG1 was found in 127 women (10.3%), G2 was found in 18 women (1.5%), and no cases of G3 were observed. Advanced Grannum grading was significantly associated with IUGR (48 [4.4%] in G0, 9 [7.1%] in G1, 5 [27.8%] in G2; P < 0.001), but was dependent on smoking status. IUGR was not predicted by abnormal MSS or abnormal UtAD findings at either the second or third trimester ultrasounds. ConclusionG2 maturation at 30 to 34 weeks’ gestation is associated with mild IUGR at delivery in low-risk women and with smoking. IUGR was not predicted by either second or third trimester markers of severe placental dysfunction. Future studies directly observing the placenta in the late third trimester may aid the elusive diagnosis of “late-onset” mild IUGR.

Placental HtrA3 Is Regulated by Oxygen Tension and Serum Levels Are Altered during Early Pregnancy in Women Destined to Develop Preeclampsia

The pathogenic origin of preeclampsia is defective placental development (placentation) and function. Preeclampsia is not diagnosed until later in pregnancy, and reliable early detection is highly desirable. HtrA3 is a recently cloned gene with high expression during placentation in the mouse, rhesus monkey, and human. The present study examined the placental production and the serum profile of HtrA3 across gestation in women, the potential molecular mechanisms regulating HtrA3 production, and the association between maternal HtrA3 serum levels and preeclampsia. Immunohistochemistry determined HtrA3 expression pattern and cellular localization in first-, second-, and third-trimester placenta. The maternal serum HtrA3 levels were analyzed by Western blotting. The regulation of placental HtrA3 production and the secretion by oxygen tension was investigated in first-trimester placental explants and trophoblast cells. Placental HtrA3 protein was maximally produced in the first trimester and then dramatically down-regulated, especially in the syncytiotrophoblast. HtrA3 was secreted into the maternal circulation with a serum profile reflecting placental production. Oxygen tension regulated HtrA3; low oxygen enhanced, whereas the transition from low to high oxygen decreased, HtrA3 protein production in syncytiotrophoblast. Maternal serum HtrA3 levels at approximately 13-14 wk of gestation were significantly higher in women who subsequently developed preeclampsia. It appeared that HtrA3 down-regulation was delayed in preeclamptic pregnancies. HtrA3 protein production is closely associated with changing in oxygen tension in the placenta. The decline in HtrA3 at the end of first trimester may reflect the placental low to high oxygen switch. Abnormally high levels of serum HtrA3 at approximately 13-14 wk of gestation is associated with preeclampsia.

Involvement of heparanase in vaginal and cesarean section deliveries

Background Heparanase is an endo-β-D-glucuronidase dominantly involved in cell invasion, tumor angiogenesis and metastasis. Recently, we demonstrated increased levels of heparanase, tissue factor pathway inhibitor (TFPI)-2 and vascular endothelial growth factor (VEGF)-A in early miscarriages (Nadir et al., Thromb Res, 2010). Herein, we investigated the role of heparanase in third trimester placentas, in correlation to tissue factor (TF), TFPI, TFPI-2, and VEGF-A. Methods Twenty five third-trimester placenta samples (weeks 36-41) were studied applying real time RTPCR and immunostaining. Ten cases were placentas of elective cesarean sections, 8 cases were of normal vaginal deliveries and 7 samples were placentas of intra-uterine growth restriction (IUGR) fetuses. Skin and lung tissues of heparanase over-expressing mice and heparanase knock-out mice were subjected to immunostaining. Results Sections obtained from vaginal and IUGR placentas revealed 2 folds increased levels of heparanase, TFPI-2 and VEGF-A compared to placentas from elective cesarean sections in maternal decidua as well as in fetal placenta elements. Interestingly, abundance of TFPI staining in the intra-villous blood was observed in placentas of vaginal and IUGR deliveries. Heparanase effect on TFPI release to the blood was supported by immunostaining of heparanase over-expressing and heparanase knock-out mice tissues. Conclusions In regard to our previous data on early pregnancy losses, the present data strengthen the understanding that in placental vascular complications levels of heparanase, TFPI-2 and VEGF-A increase. In the process of delivery, heparanase may have a regulatory role on TFPI release to fetal circulation.

Strip of the Month: July 2010

Strip of the Month: July 2010