Early Placentation Research Articles

In Vitro Culture of Camelid Embryos.

The camel is the main meat- and milk-producing animal in the desert environment and is characterized by an induced ovulation pattern, in which ovulation occurs in response to copulation. Little is known about the early embryonic development and placentation in camelid species. Here we describe protocols for the culture of both in vitro-produced and in vivo-retrieved camel embryos. A chemically defined medium enables the development of in vitro-produced embryos from cleavage to the hatching blastocyst stage. In vivo-retrieved embryos will survive in vitro for 23days postinsemination, reaching a diameter of ~5mm.

Inflammatory mediators: a causal link to hypertension during preeclampsia.

Preeclampsia (PE) is a hypertensive disorder that occurs after 20weeks of gestation, implicating the placenta as a key offender. PE is associated with an imbalance among B lymphocytes, CD4+ T lymphocytes, NK cells and increased inflammatory cytokines. During early onset PE, trophoblast invasion and placentation are impaired, leading to reduced blood flow to the fetus. In all spectrums of this disorder, a shift towards a pro-inflammatory state where regulatory cells and cytokines are decreased occurs. Specifically, inflammatory CD4+ T-cells and inflammatory cytokines are increased while CD4+ T regulatory cells (Tregs) and immunosuppressive cytokines such as IL-4 and IL-10 are decreased resulting in B cell activation, production of autoantibodies, endothelial dysfunction and hypertension associated with PE. However, the stimulus for these imbalances is unknown and need to be fully understood so that effective treatments that target the pathogenesis of the disease can be designed. Therefore, this review will focus on the pathways involving CD4+ , TH1, TH2, Tregs, TH17s, B cells, and NK cells in the pathophysiology of PE. LINKED ARTICLES: This article is part of a themed section on Immune Targets in Hypertension. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.12/issuetoc.

97. Effect of placental growth factor on trophoblast integration into endothelial cell networks in the presence of inflammation

Introduction Invasion of maternal spiral arterioles by trophoblast cells is fundamental to normal placental development. Abnormal spiral arteriole remodelling is often present in pregnancies affected by preeclampsia. Placental growth factor (PlGF) is produced by the placenta and this molecule is decreased during early pregnancy of women with preeclamptic pregnancy as compared to unaffected women. The influence of PlGF on the processes of early placentation is not known. Objectives To observe the effect of supplemental PlGF upon capillary-like uterine endothelial cell networks and a first trimester human trophoblast cell line in the presence or absence of inflammation induced by exogenous tumour necrosis factor-alpha (TNF- α ) . Methods 24-well tissue culture plates were coated with 300 uL of undiluted Matrigel and allowed to gelatinise at 37°C for 30 min. Fluorescent labelled uterine microvascular endothelial cells and HTR8/SVNeo cells were co-cultured (1 X 10 5 per well) for 20 h treated with PlGF (10 ng/mL) and/or TNF- α (0.5 ng/mL). Images were captured by fluorescence microscopy and analysed using ImageJ. Experiments were repeated 8 times, data was analysed using SPSS v24. Results TNF alpha reduced trophoblast cell integration into endothelial networks (Control vs TNF- α p = 0.006) but this was not ameliorated by addition of PlGF (TNF- α vs TNF- α + PlGF p > 0.9). Furthermore, PlGF supplementation itself did not improve cell integration (Control vs PlGF p = 0.2). Discussion PlGF not improve trophoblast cell integration into endothelial cell networks nor reverse the inhibitory effect of TNF- α .

Placental formation in early pregnancy: how is the centre of the placenta made?

Correct development of the placenta is critical to establishing pregnancy and inadequate placentation leads to implantation failure and miscarriage, as well as later gestation pregnancy disorders. Much attention has been focused on the placental trophoblasts and it is clear that the trophoblast lineages arise from the trophectoderm of the blastocyst. In contrast, the cells of the placental mesenchyme are thought to arise from the inner cell mass, but the details of this process are limited. Due to ethical constraints and the inaccessibility of very early implantation tissues, our knowledge of early placentation has been largely based on historical histological sections. More recently, stem cell technologies have begun to shed important new light on the origins of the placental mesenchymal lineages. This review aims to amalgamate the older and more modern literature regarding the origins of the non-trophoblast lineages of the human placenta. We highlight ways in which rapidly developing stem cell technologies may shed new light on these crucial peri-implantation events. Relevant articles were identified using the PubMed database and Google Scholar search engines. A pearl growing method was used to expand the scope of papers relevant to the cell differentiation events of non-trophoblast placental lineages. At the start of pregnancy, cells of the extraembyronic mesoderm migrate to underlie the primitive trophoblast layers forming the first placental villi. The mesenchymal cells in the villus core most likely originate from the hypoblast of the embryo, but whether cells from the epiblast also contribute is yet to be determined. This is important because, following the formation of the villus core, vasculogenesis and haematopoiesis take place in the nascent placenta before it is connected to the embryonic circulation, making it likely that haematopoietic foci, placental macrophages, endothelial cells and vascular smooth muscle cells all arise in the placenta de novo. Evidence from the stem cell field indicates that these cells could directly differentiate from the extraembryonic mesoderm. However, the lineage hierarchy involved in cell fate decisions has not been well-established. Mesodermal progenitors capable of differentiating into both vascular and haematopoietic lineages can be derived from human embryonic stem cells, but the identification of such stem cells in the placenta is lacking. Future work profiling rare progenitor populations in early placentae will aid our understanding of early placentation. Understanding the origins of the cell lineages of the normal placenta will help us understand why so many pregnancies fail and address mechanisms that may salvage some of these losses.

Epigenetic changes in preterm birth placenta suggest a role for ADAMTS genes in spontaneous preterm birth.

Preterm birth (PTB) affects approximately 1 in 10 pregnancies and contributes to approximately 50% of neonatal mortality. However, despite decades of research, little is understood about the etiology of PTB, likely due to the multifactorial nature of the disease. In this study, we examined preterm and term placentas, from unassisted conceptions and those conceived using in vitro fertilization (IVF). IVF increases the risk of PTB and causes epigenetic change in the placenta and fetus; therefore, we utilized these patients as a unique population with a potential common etiology. We investigated genome-wide DNA methylation in placentas from term IVF, preterm IVF, term control (unassisted conception) and preterm control pregnancies and discovered epigenetic dysregulation of multiple genes involved in cell migration, including members of the ADAMTS family, ADAMTS12 and ADAMTS16. These genes function in extracellular matrix regulation and tumor cell invasion, processes replicated by invasive trophoblasts (extravillous trophoblasts (EVTs)) during early placentation. Though expression was similar between term and preterm placentas, we found that both genes demonstrate high expression in first- and second-trimester placenta, specifically in EVTs and syncytiotrophoblasts. When we knocked down ADAMTS12 or ADAMTS16in vitro, there was poor EVT invasion and reduced matrix metalloproteinase activity, reinforcing their critical role in placentation. In conclusion, utilizing a population at high risk for PTB, we have identified a role for ADAMTS gene methylation in regulating early placentation and susceptibility to PTB.

Excess glucose induce trophoblast inflammation and limit cell migration through HMGB1 activation of Toll-Like receptor 4.

Hyperglycemia increases the risk of preeclampsia. Hyperglycemia induces a placental trophoblast inflammatory (IL-1β, IL-6, IL-8), antiangiogenic (sFlt-1, sEndoglin), and anti-migratory profile. The IL-1β response is mediated via inflammasome activation by the damage-associated molecular pattern (DAMP), uric acid. The objective of this study was to determine the role of high-mobility group box-1 (HMGB1), a DAMP that activates Toll-like receptor 4 (TLR4), in human first trimester trophoblast responses to hyperglycemia. The trophoblast response to excess glucose under different oxygen tensions was also investigated. The human first trimester trophoblast cell line (Sw.71) was exposed to glucose mimicking normoglycemia (5mmol/L) and hyperglycemia (10mmol/L), either alone or with the TLR4 antagonist, LPS-RS; or the HMGB1 inhibitor, glycyrrhizin. Cells were also treated with glucose under hyperoxic (21% O2 ), normoxic (8% O2 ), and hypoxic (2% O2 ) conditions. Cell-free supernatants were assayed by ELISA and bioassays for inflammatory: IL-1β, IL-6, and IL-8; inflammasome-associated: uric acid and caspase-1; angiogenic: sEndoglin, sFlt-1, and PlGF; and the DAMP, HMGB1. Cell migration was measured using a two-chamber colormetric assay. Excess glucose triggered a trophoblast sterile inflammatory IL-8 and antimigratory response through HMGB1 activation of TLR4. The IL-1β and sFlt-1/PlGF response was TLR4-mediated, but HMGB1-independent, suggesting another DAMP may be involved. Hyperoxia rather than normoxia or hypoxia was a major driver of trophoblast dysfunction in response to excess glucose. The findings from this study indicate a novel mechanism by which hyperglycemia may impact trophoblast function, early placentation, and ultimately, pregnancy outcome.

Trichlorfon inhibits proliferation and promotes apoptosis of porcine trophectoderm and uterine luminal epithelial cells

Trichlorfon is an organophosphate insecticide widely used in agriculture. Additionally, it is applied to pigs for control of endo- and ectoparasites. Previous studies have shown the effects of trichlorfon in pigs during late stages of gestation; however, little is known about its effects during early pregnancy, including implantation and placentation. We investigated whether trichlorfon affects proliferation and apoptosis of porcine trophectoderm (pTr) and uterine luminal epithelial (pLE) cells. Trichlorfon inhibited the proliferation of pTr and pLE cells, as evidenced by cell cycle arrest, and altered the expression of proliferation-related proteins. In addition, trichlorfon induced cell death and apoptotic features, such as loss of mitochondrial membrane potential and DNA fragmentation, in pTr and pLE cells. Moreover, trichlorfon treatment decreased concentrations of Ca2+ in the cytoplasm in both cell lines and increased concentrations of Ca2+ in mitochondria of pTr cells. Trichlorfon inhibited the activation of phosphoinositide 3-kinase/AKT and mitogen-activated protein kinase signaling pathways in pTr and pLE cells. Therefore, we suggest that trichlorfon-treated pTr and pLE cells exhibited abnormal cell physiology which might lead to early pregnancy failure.

Second trimester maternal serum–amniotic fluid nitric oxide and vascular endothelial growth factor levels in relation to uterine artery Doppler indices in pregnancies with normal outcome

The aim of the present study was to explore the possible relations of maternal serum and amniotic fluid nitric oxide (NO) and of vascular endothelial growth factor A (VEGF-A) concentrations with uterine artery Doppler indices (UtADs) in a healthy second trimester obstetric population. In this regard, the levels of NO and VEGF-A were measured in maternal sera and the amniotic fluid samples were in 36 subjects who were in their early second trimester of pregnancy. The mean levels for VEGF-A were 110.3 ± 31.5 pg/ml in maternal serum and 149.6 ± 31.3 pg/ml in amniotic fluid. Mean levels for NO were 5.7 ± 4.7 µmol in maternal serum and 12.9 ± 3.4 µmol in amniotic fluid. UtADs were measured concurrently with the sample collections. The mean value for uterine artery pulsatility index (PI) was 1.3 ± 0.4. The measurements were then analysed for possible correlations, whereby no correlation was found between UtAD and maternal serum levels of either molecule (p = .828 and p = .662 for VEGF-A and NO, respectively). However, a positive correlation was found between the NO levels in the amniotic fluid compartment and UtAD (r = 0.432, p = .009 for PI). Therefore, a correlation of UtAD with amniotic fluid NO can be expected in pregnancies with a normal outcome.Impact StatementWhat is already known on this subject? Nitric oxide (NO) and vascular endothelial growth factor A (VEGF-A) are important vasoactive molecules that play significant roles in early angiogenesis and placentation.What the results of this study add? There is a positive correlation between the amniotic fluid NO levels and the uterine artery Doppler indices (UtADs) in the second trimester of pregnancies with normal outcomes.What the implications are of these findings for clinical practice and/or future research? A correlation between amniotic fluid NO levels and UtAD may indicate a normal trophoblastic invasion. Disturbance of this balance may be expected in certain adverse pregnancy outcomes. Additional studies are needed to further explore the molecular signs of early abnormal placentation and their clinical reflection.

Placenta accreta spectrum: a need for more research on its aetiopathogenesis.

Placenta accreta is a pathologic condition of early placentation that has been studied by clinicians and pathologists for nearly 100 years. The main effect is a loss of the normal plane of cleavage from the uterine wall, resulting in massive haemorrhage when attempts are made at removing the placenta manually at delivery. Excessive remodelling of the radial and arcuate arteries in the invasive forms of placenta accreta spectrum increases the risk of complications (Jauniaux et al., AJOG, 2018;218:75-87).

SUPPLERERMATIC PICTURE OF EXTREMIBLE BLOOD IN THE EARLY THREE-MONTH OF DOOPLER IMAGE OF EXTRA-EMBRYONIC BLOOD FLOW AT THE EARLY GESTATION TRIMESTER AGAINST THE GROUND OF INFLAMMATORY DISEASES OF THE FEMALE REPRODUCTIVE ORGANS

One of the main complications during early embryogenesis and placentation is placental dysfunction. Under the influence of various factors, namely infectious ones, which depending on biological specificity negatively affect the reproductive cells of parents, the development of the embryo, the formation of trophoblast and placenta, which in turn leads to disorders of the placental complex function and gestational complications during pregnancy. Due to the expansion of diagnostic capabilities for detecting disorders of the placenta function, as well as the emergence of new data on the mechanisms of blood flow regulation in the placenta to evaluate the functional state of the feto-placental complex (FPC) at the phase of the primary placental dysfunction formation in women with pelvic inflammatory diseases, we identified Doppler image of the blood flow features (ovarian artery, inter-villous space) in women of the main and control groups. According to the objective, an ultrasound examination of 67 women in 6-7 and 12-14 weeks of gestation was performed. The main group included 37 women with signs of inflammatory diseases of the female genital organs, and the control group included 30 practically healthy women. Obtaining new data on the mechanisms of blood circulation regulation in the ovarian artery and inter-villous space at the stage of placental complex formation during physiological and complicated pregnancy will enable to supplement the issues of pathogenesis of development and prevention of placental dysfunction.

Recurrent Implantation Failure and Low Molecular Weight Heparin

Implantation of the embryo into the endometrium is the first step in the establishment of pregnancy. This process is complex, and depends on many factors. Recurrent implantation failure is a source of distress to patients and specialists. It is defined as failure to achieve a viable pregnancy, following “>3 embryo transfers with high quality embryos or the transfer of ≥ 10 embryos in multiple transfers”. Thrombophilic conditions that contribute to recurrent implantation failure are the main concern in this review. The mechanism of implantation failure is believed to be due to decreased blood flow to the endometrium and placenta which can hinder normal endometrial receptivity leading to miscarriage. Defects in early placentation resulting in pregnancy failure, have focused attention on the therapeutic potential of low molecular weight heparin in the implantation process. Heparin has a role at all stages of implantation to improve pregnancy outcomes. There are controversies in literature regarding the association between thrombophilia and recurrent implantation failure and available literature regarding this issue is very heterogeneous. Various investigators, have shown that women with RIF are more likely to have a thrombophilia disorder, yet a clear cause cannot be acknowledged from these studies. Heparin treatment has been evaluated in several studies, showing conflicting evidence. However, several studies have pointed out that it may play a role in a subset of patients who presents a thrombophilia mutation, thus the group of patients that might benefit is needed to be identified. This review is dedicated to evaluate the published literature about the role of low molecular weight heparin in case of recurrent implantation failure with or without the presence of thrombophilia.

Chronically elevated androgen with or without a western-style diet reduces the pregnancy rate and early placental vascularization in young adult rhesus monkeys

To determine if chronic hyperandrogenemia and/or consumption of a western-style diet (WSD) beginning near menarche suppresses pregnancy rate or early placentation in young adults. 2 by 2 factorial cohort Silastic implants containing either cholesterol (control C, n=20) or T (serum T levels ∼1.5 ng/ml; n=20) were placed sc in rhesus macaque females nearing menarche. These females were randomized to consume either a standard chow diet, or a WSD in a 2 x 2 factorial experimental design (n=9-10/group). After 3.5 years of treatment (∼6 years old), individuals were paired with a fertile male at the mid-cycle rise in serum E2 in a time-mated breeding (TMB) program [1]. Serum P4 was monitored post-TMB until levels either: a) declined below 0.5 ng/ml with concurrent menses, i.e., pregnancy did not occur, or b) remained elevated. Pregnancies were confirmed by Doppler ultrasound (DUS) on gestational day (GD) 30-36, and placental blood volume (BV) determined by contrast-enhanced US [1]. Pregnancy rates and number of TMB to pregnancy were analyzed by Generalized Linear Model function (chi-squared values), and placental BV was analyzed by Mixed Models function of SAS. Thirty-eight monkeys fulfilled the criteria for mating; 1 WSD-treated female was acyclic and did not display a rise in serum E2 levels. Similar numbers of the control (C, 7/10; 70%) and WSD (7/10; 70%) females became pregnant, whereas fewer T (4/10; 40%) and T+WSD (3/9; 33%) females achieved pregnancy (effect of steroid P<0.05) at first pairing. Also, 2 of 3 pregnancies in the T+WSD group were atypical: 1 anembryonic pregnancy miscarried by GD58-59, and 1 twin pregnancy was delivered early at GD105 due to pregnancy complications. Additional TMB (up to 3x) resulted in 100% of C, 70% of WSD, 100% of T and 69% of T+WSD females conceiving (diet P<0.004), however T-treated females required more TMB to achieve pregnancy than C or WSD groups (steroid P<0.02). At GD30-36, the BV of placentas in T, WSD and T+WSD females was reduced 1.6 (T), 2.1 (WSD), and 1.8 (T+WSD)-fold compared to controls (P<0.05). Exposure to chronically elevated androgen significantly reduced pregnancy success in young adult females at first mating with fertile male monkeys, and delayed time to pregnancy. Based on our previous studies, the subfertility could be due to: a) ovulation of immature or degenerated oocytes, or b) abnormal expression of endometrial decidualization and implantation markers detected at mid-secretory phase of the menstrual cycle in T and T+WSD treated animals. Reduced placental BV during the first trimester in T, WSD and T+WSD-treated females, may impair fetal-placental health as gestation progresses, which is currently under investigation. These data may provide insight into the etiology of subfertility in women experiencing hyperandrogenemia despite continued ovarian cyclicity.

Hypoxia-induced TET1 facilitates trophoblast cell migration and invasion through HIF1\u03b1 signaling pathway

Low oxygen is a typical extrinsic factor for the regulation of trophoblast biological function, including cell migration, invasion and proliferation. Ten-eleven translocation methylcytosine dioxygenase 1 (TET1), an enzyme converting 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC), is transcriptionally activated by hypoxia in cancer cells. Therefore, we focus on the role of TET1 on trophoblast function in a physiologically hypoxic environment (3% oxygen), which is related to early placentation. Here, we found that TET1 was highly expressed in first trimester villi compared with normal term placentas. In vitro, both TET1 mRNA and protein expression levels in JEG3 cells were increased following exposure to 3% oxygen, and the migration and invasion capacities of JEG3 cells were up-regulated. Furthermore, TET1 knockdown decreased the migration, invasion and proliferation of JEG3 cells exposed to 3% oxygen, and the expression of HIF1α and its downstream target genes was also decreased, which was related to hyper-methylation of the HIF1α promoter. Finally, increased HIF1α protein expression reversed the inhibitory effect of TET1 knockdown on the migration and invasion of JEG3 cells exposed to 3% oxygen. These data show that hypoxia-induced TET1 expression facilitates trophoblast cell migration and invasion through the HIF1α signaling pathway, which plays an important role during placentation.

Isolation and characterization of the trophectoderm from the Arabian camel (Camelus dromedarius)

We isolated and characterized trophoblast from in vivo-derived camel embryos and compared with embryonic stem-like cells. Camel embryos were flushed on day 8 post-insemination and used to derive trophectoderm and embryonic stem-like cells under feeder-free culture conditions using a basement membrane matrix. Embryos were evaluated for the expression of POU5F1, MYC, KLF4, SOX2, CDX2, and KRT8 mRNA transcripts by relative quantitative polymerase chain reaction. Camel embryos grew and expanded to ∼4.5 mm and maintained their vesicular shape in vitro for 21 days post-insemination. Trophoblast and embryonic stem-like cell lines grew under feeder-free culture conditions and showed distinct morphological criteria and normal chromosomal counts. Embryonic stem-like cells showed positive staining in the alkaline phosphatase reaction. Trophoblast cells showed a significant increase in CDX2, KRT8, KLF4, and SOX2 expression compared with embryonic stem-like cells and whole embryos. Embryonic stem-like cells showed a significant decrease in CDX2 expression and increase in SOX2 and KRT8 expression compared to embryonic expression. POU5F1 and MYC expression showed no difference between embryos and both cell lines. We characterized embryo survival in vitro, particularly the derivation of trophectoderm and embryonic stem-like cells, providing a foundation for further analysis of early embryonic development and placentation in camels.

146 Free fetal haemoglobin impacts fetoplacental vascular structure and function: implications for fetal growth restriction

IntroductionIn fetal growth restriction (FGR), early placentation defects result in placental insufficiency and vasoconstriction, which cumulate in organ failure at the severe end of the spectrum. In other placental pathologies,...

Extravillous Trophoblast and Endothelial Cell Crosstalk Mediates Leukocyte Infiltration to the Early Remodeling Decidual Spiral Arteriole Wall.

Decidual spiral arteriole (SpA) remodeling is essential to ensure optimal uteroplacental blood flow during human pregnancy, yet very little is known about the regulatory mechanisms. Uterine decidual NK (dNK) cells and macrophages infiltrate the SpAs and are proposed to initiate remodeling before colonization by extravillous trophoblasts (EVTs); however, the trigger for their infiltration is unknown. Using human first trimester placenta, decidua, primary dNK cells, and macrophages, we tested the hypothesis that EVTs activate SpA endothelial cells to secrete chemokines that have the potential to recruit maternal immune cells into SpAs. Gene array, real-time PCR, and ELISA analyses showed that treatment of endothelial cells with EVT conditioned medium significantly increased production of two chemokines, CCL14 and CXCL6. CCL14 induced chemotaxis of both dNK cells and decidual macrophages, whereas CXCL6 also induced dNK cell migration. Analysis of the decidua basalis from early pregnancy demonstrated expression of CCL14 and CXCL6 by endothelial cells in remodeling SpAs, and their cognate receptors are present in both dNK cells and macrophages. Neutralization studies identified IL-6 and CXCL8 as factors secreted by EVTs that induce endothelial cell CCL14 and CXCL6 expression. This study has identified intricate crosstalk between EVTs, SpA cells, and decidual immune cells that governs their recruitment to SpAs in the early stages of remodeling and has identified potential key candidate factors involved. This provides a new understanding of the interactions between maternal and fetal cells during early placentation and highlights novel avenues for research to understand defective SpA remodeling and consequent pregnancy pathology.

Dynamic maternal and fetal Notch activity and expression in placentation

IntroductionMurine placentation requires trophoblast Notch2, while the Notch ligand, JAGGED1, is reduced in invasive trophoblasts from women with preeclampsia. However, the placental cells with active Notch signaling and expression of other Notch proteins and ligands in placentation have yet to be defined. We sought to identify endothelial cell and trophoblast subtypes with canonical Notch signaling in the decidua and placenta and correlate this to expression of Notch proteins and ligands. MethodsNotch reporter transgenic mice were used to define canonical Notch activity and immunofluorescence staining performed to characterize expression of Notch1, 2, 3, 4 and ligands, Delta-like 4 (Dll4) and Jagged1 (Jag1) during early placentation and in the mature placenta. ResultsNotch signaling is active in maternal and fetal endothelial cells and trophoblasts during early placentation and in the mature placenta. Dll4, Jag1, Notch1, and Notch4 are expressed in maternal vasculature in the decidua. Dll4, Jag1 and Notch1 are expressed in fetal vasculature in the labyrinth. Dll4, Notch2 and Notch4 are co-expressed in the ectoplacental cone. Notch2 and Notch4 are expressed in parietal-trophoblast giant cells and junctional zone trophoblasts with active canonical Notch signaling and in labyrinthine syncytiotrophoblasts and sinusoidal-trophoblast giant cells. DiscussionCanonical Notch activity and distinct expression patterns for Notch proteins and ligands was evident in endothelium and trophoblasts, suggesting Notch1, Notch2, Notch4, Dll4, and Jag1 have distinct and overlapping functions in placentation. Characterization of Notch signaling defects in existing mouse models of preeclampsia may shed light on the role of Notch in developing the preeclampsia phenotype.

Effect of proline rich 15-deficiency on trophoblast viability and survival.

Deviations from the normal program of gene expression during early pregnancy can lead to early embryonic loss as well as dysfunctional placentation, which can cause significant morbidity and mortality. Proline rich 15 (PRR15) is a low molecular weight nuclear protein expressed by the trophoblast during early gestation. Lentivirus-mediated knockdown of PRR15 mRNA in ovine trophectoderm led to demise of the embryo by gestational day 15, providing compelling evidence that PRR15 expression is critical during this precarious window of development. Our objective was to determine the effect of PRR15 knockdown on trophoblast gene expression, proliferation, and survival. The first-trimester human trophoblast cell line, ACH-3P, was infected with control lentivirus or a lentivirus expressing a short hairpin (sh)RNA to target PRR15 mRNA for degradation, resulting in a 68% reduction in PRR15 mRNA. Microarray analysis of these cell lines revealed differential expression of genes related to cancer, focal adhesion, and p53 signaling. These changes included significant up-regulation of GDF15, a cytokine increased in pregnancies with preeclampsia. Viability and proliferation decreased in PRR15-deficient cells, which was consistent with down-regulation of cell cycle-related genes CCND1 and CDK6 and an up-regulation of CCNG2 and CDKN1A in the PRR15-deficient cells. TNFSF10, a tumor necrosis factor superfamily member known to induce apoptosis increased significantly in the PRR15-deficient cells. Migration through a basement membrane matrix decreased and an increased population of apoptotic cells was present when treated with shRNA to target PRR15. These results suggest that PRR15 enhances trophoblast viability and survival during early implantation and placentation.

Regulation of the prorenin - angiotensin system by oxygen and miRNAs; parallels between placentation and tumour development?

Tissue renin-angiotensin systems (RASs) are involved in tissue growth and development as they are important regulators of angiogenesis, cell proliferation and migration. The placental RAS is most highly expressed in early gestation, at a time when the oxygen tension within the conceptus is reduced, and plays a key role in placental growth and development. Similar to the placenta, tumour development relies on proliferation, angiogenesis and invasion in order to grow and metastasize. The RAS is known to be upregulated in a variety of solid tumours, including ovarian, endometrial, cervical, breast and prostate. This review explores the roles of oxygen and microRNAs in regulating the normal expression of the placental RAS, providing insight into regulation of its development as well as the development of disease states in which the RAS is overexpressed. We propose that the placental RAS is downregulated by microRNAs that are suppressed during the physiologically normal ‘hypoxic’ phase of early placentation. Suppression of these miRNAs allows the placental RAS to stimulate placental growth and angiogenesis. We propose that similar mechanisms may be at play in solid tumours, which are characterised by hypoxia.

Androgen dependent mechanisms of pro-angiogenic networks in placental and tumor development

The placenta and tumors share important characteristics, including a requirement to establish effective angiogenesis. In the case of the placenta, optimal angiogenesis is required to sustain the blood flow required to maintain a successful pregnancy, whereas in tumors establishing new blood supplies is considered a key step in supporting metastases. Therefore the development of novel angiogenesis inhibitors has been an area of active research in oncology. A subset of the molecular processes regulating angiogenesis are well understood in the context of both early placentation and tumorigenesis. In this review we focus on the well-established role of androgen regulation of angiogenesis in cancer and relate these mechanisms to placental angiogenesis. The physiological actions of androgens are mediated by the androgen receptor (AR), a ligand dependent transcription factor. Androgens and the AR are essential for normal male embryonic development, puberty and lifelong health. Defects in androgen signalling are associated with a diverse range of clinical disorders in men and women including disorders of sex development (DSD), polycystic ovary syndrome in women and many cancers. We summarize the diverse molecular mechanisms of androgen regulation of angiogenesis and infer the potential significance of these pathways to normal and pathogenic placental function. Finally, we offer potential research applications of androgen-targeting molecules developed to treat cancer as investigative tools to help further delineate the role of androgen signalling in placental function and maternal and offspring health in animal models.