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The aim of this paper is to review available data about drugs for preventing preterm labour. Tocolytic therapy includes β adrenergic receptor agonists, NO donors, magnesium sulphate, prostaglandin-synthase inhibitors, oxytocin receptor antagonists, calcium-channel blockers, progesterone, 17-α-hydroxyprogesterone caproate, and antibiotics. Their specific effects on myometrial contractility, their safety, their efficiency, and side effects profile for the mother and the fetus are presented. The main question of why and for what reasons tocolysis should be administrated is discussed.

Preeclampsia is a severe complication of pregnancy, affecting an estimated 4 million women annually. It is one of the leading causes of maternal and fetal mortality worldwide, and it has life-long consequences. The maternal multisystemic symptoms are driven by poor placentation, which causes syncytiotrophoblastic stress and the release of factors into the maternal bloodstream. Amongst them, the soluble fms-like tyrosine kinase-1 (sFLT-1) triggers extensive endothelial dysfunction by acting as a decoy receptor for the vascular endothelial growth factor (VEGF) and the placental growth factor (PGF). Current interventions aim to mitigate hypertension and seizures, but the only definite treatment remains induced delivery. Thus, there is a pressing need for novel therapies to remedy this situation. Notably, CBP-4888, a siRNA drug delivered subcutaneously to knock down sFLT1 expression in the placenta, has recently obtained Fast Track approval from the Food and Drug Administration (FDA) and is undergoing a phase 1 clinical trial. Such advance highlights a growing interest and significant potential in gene therapy to manage preeclampsia. This review summarizes the advances and prospects of gene therapy in treating placental dysfunction and illustrates crucial challenges and considerations for these emerging treatments.

Background: Preeclampsia is one of the leading causes of maternal mortality worldwide and is strongly associated with long-term morbidity in mothers and newborns. Referred to as one of the deep placentation disorders, insufficient remodeling of the spiral arteries during the first trimester remains a major cause of placental dysfunction. Persisting pulsatile uterine blood flow causes abnormal ischemia/reoxygenation phenomenon in the placenta and stabilizes the HIF-2α (hypoxia-inducible factor-2α) in the cytotrophoblasts. HIF-2α signaling impairs trophoblast differentiation and increases sFLT-1 (soluble fms-like tyrosine kinase-1) secretion, which reduces fetal growth and causes maternal symptoms. This study aims to evaluate the benefits of using PT2385-an oral specific HIF-2α inhibitor-to treat severe placental dysfunction., Methods: To evaluate its therapeutic potential, PT2385 was first studied in primary human cytotrophoblasts isolated from term placenta and exposed to 2.5% O 2 to stabilize HIF-2α. Viability and luciferase assays, RNA sequencing, and immunostaining were used to analyze differentiation and angiogenic factor balance. The ability of PT2385 to mitigate maternal manifestations of preeclampsia was studied in the selective reduced uterine perfusion pressure model performed in Sprague-Dawley rats., Results: In vitro, RNA sequencing analysis and conventional techniques showed that treated cytotrophoblast displayed an enhanced differentiation into syncytiotrophoblasts and normalized angiogenic factor secretion compared with vehicle-treated cells. In the selective reduced uterine perfusion pressure model, PT2385 efficiently decreased sFLT-1 production, thus preventing the onset of hypertension and proteinuria in pregnant dams., Conclusions: These results highlight HIF-2α as a new player in our understanding of placental dysfunction and support the use of PT2385 to treat severe preeclampsia in humans.

Background: Pregnancies in women who underwent kidney transplants are at high risk compared with the general population., Methods: In this study, we aimed to retrospectively assess the obstetrical complications, delivery outcomes, and impact of pregnancy on kidney allograft function in a single-center cohort of kidney transplant recipients (KTRs). We provide data regarding the long-term evolution of children., Results: Thirty-two KTRs underwent a total of 57 pregnancies between 1994 and 2010. Fourteen pregnancies (24 %) did not survive caused by miscarriages (n = 9), stillborn (n = 1), ectopic pregnancies (n = 2), and medical abortion (n = 2). Live birth occurred in 76% of pregnancies. Delivery was by cesarean in 66%. The mean gestational age was 30.45 ± 11.3 weeks and 65% of newborns were premature. A low birth weight <2500g was noted in 46%. Obstetric complications were de novo hypertension in 4%, pre-eclampsia in 9%, and gestational diabetes in 2%. The 5- and 10-year post-delivery death-censored graft loss rates were 3.1% and 12.5%, respectively. Data on 21 children were collected via a self-questionnaire. After a median follow-up time of 17 years, they appeared in good medical and psychological health. None of them suffered from chronic disease (especially uronephrological condition) or was taking chronic medication., Conclusions: Long-term evolution of children born to women who underwent kidney transplants seems favorable. Pregnancies in KTRs are successful in two-thirds of cases but are at increased risk of prematurity, delivery by cesarean, and low birth weight., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The placenta is a multifaceted organ, fulfilling critical functions for the fetus and the mother. Therefore, it is a critical regulator of the pregnancy, and its dysfunction leads to diseases, including fetal growth restriction and preeclampsia. Studying the placenta is a difficult task since its existence is transient, and its structure is specific to our species. In vitro differentiation of primary cytotrophoblast isolated from term human placenta has been widely used in the placental research field as it represents a reliable model to study cellular differentiation and function. Direct alternatives include trophoblastic cell lines, explants, and organoids, but this protocol, based on the separation of the cells on a Percoll gradient, presents the advantage of being relatively cheap and easy to perform in every research laboratory. Furthermore, the 2D culture is a flexible method that can be adapted to various experimental conditions (transfection, drug exposure, metabolic study, observations, etc .), allowing mechanistic explorations of cellular processes., (Copyright © 2021 The Authors; exclusive licensee Bio-protocol LLC.)

Introduction : A loss of endogenous stem cells capable of tissue repair and regeneration drives the biological process that we recognize as "aging". Recovery of stem cell-mediated repair and regenerative functions in aged animals has been reported in murine heterochronic parabiosis experiments. Objectives : Herein we will review how pregnancy is an unusual form of heterochronic parabiosis, as the placenta prevents the exchange of most blood cells between parabionts. Instead, plasma and its content, including small extracellular vesicles, can readily cross the placental barrier. These nanosized extracellular vesicles are readily produced by the placenta, amnion, fetus and mother, and are essential for fetal organogenesis, growth and the progression of a healthy pregnancy. If defective, these extracellular vesicles can cause havoc such as in the case of peripartum cardiomyopathy. We will also review how these extracellular vesicles impact the mother substantially (including cardiac function) in the parabiosis of pregnancy. Conclusion : Extracellular vesicles generated during the course of a healthy pregnancy are essential for organogenesis and fetal growth, and also for maternal tissue repair and regeneration, and might be defective or deficient in pregnancies that result in peripartum cardiomyopathy., (© 2021 The Author(s). Published by BRI.)

Despite occasional reports of vertical transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) during pregnancy, the question of placental infection and its consequences for the newborn remain unanswered. Herein, we analyzed the placentas of 31 coronavirus disease 2019-positive mothers by reverse transcriptase PCR, immunohistochemistry, and in situ hybridization. Only one case of placental infection was detected, which was associated with intrauterine demise of the fetus. Differentiated primary trophoblasts were then isolated from nonpathologic human placentas at term, differentiated, and exposed to SARS-CoV-2 virions. Unlike for positive control cells Vero E6, the virus inside cytotrophoblasts and syncytiotrophoblasts or in the supernatant 4 days after infection was undetectable. As a mechanism of defense, we hypothesized that trophoblasts at term do not express angiotensin-converting enzyme 2 and transmembrane protease serine 2 (TMPRSS2), the two main host membrane receptors for SARS-CoV-2 entry. The quantification of these proteins in the placenta during pregnancy confirmed the absence of TMPRSS2 at the surface of the syncytium. Surprisingly, a transiently induced experimental expression of TMPRSS2 did not allow the entry or replication of the virus in differentiated trophoblasts. Altogether, these results underline that trophoblasts are not likely to be infected by SARS-CoV-2 at term, but raise concern about preterm infection., (Copyright © 2021 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Insufficient remodeling of uterine arteries causes pregnancy-related diseases, including fetal growth restriction and preeclampsia. In these situations, reduced maternal blood flow in the placenta is thought to be responsible for the persistence of a low oxygen environment throughout pregnancy. We hypothesized that chronic activation of transcription factors hypoxia-inducible factors (HIFs) actively participates in placental underdevelopment, which impairs fetal growth. The computer-assisted analysis in pathological placentas revealed an increased number of HIF-2α-positive nuclei in the syncytium compared to normal human placentas, while HIF-1α stabilization was unchanged. Specific involvement of HIF-2α was confirmed in primary human cytotrophoblasts rendered deficient for HIF1A or HIF2A. Silencing HIF2A increased the expression of main syncytialization markers as well as differentiation and syncytium formation. It also improved placental growth factor bioavailability. None of these changes was seen when silencing HIF1A. Conversely, the experimental induction of HIF-2α expression repressed forskolin-induced differentiation in BeWo choriocarcinoma cells. Our mechanistic insights evidence that transcription factor HIF-2α impairs placental function, thus suggesting its participation in fetal growth restriction and preeclampsia when placentas become chronically hypoxic. Furthermore, it suggests the possibility to develop novel molecular targeting therapies for placental dysfunction., (© 2020 Federation of American Societies for Experimental Biology.)

Background: The diagnosis of cervical insufficiency is based on the previous history of recurrent second or early third trimester losses. Its incidence among pregnant women is 0.5-1% but can be as high as 75% among women with preterm birth. Transvaginal cerclage (TVC) is the common therapy of cervical insufficiency. However, this technique has several limits, especially in twin pregnancies. As some selected conditions, a transabdominal cerclage (TAC) is indicated, it has been offered to patients with multiple pregnancies. Aim: To evaluate the outcomes of twin pregnancies with transabdominal cerclage in terms of preterm birth rate and neonatal morbidity and mortality. Materials and methods: We conducted a retrospective study of seven patients with twin pregnancies managed with transabdominal cerclage at the end of the first trimester (12-15 weeks). We selected patients with a history of fetal loss who met the indications criteria of TAC (history of TVC failure or short cervix unable to have TVC). The antenatal and delivery data were collected and compared to those of their previous pregnancy. Outcomes: All patients carried their pregnancy throughout the second trimester and delivered during the third trimester. Mean gestational age was 34 4/7 week. All newborns were alive and neonatal morbidity rate was 50%, mostly related to preterm birth. Mean duration of neonatal intensive care stay was 32 days. There were no operative complications following TAC. Conclusions: Perinatal outcomes are considerably improved in twin pregnancies with transabdominal cerclage. Our findings corroborate those of previous case reports and support the efficacy of TAC for managing cervical insufficiency in twin pregnancies.

Introduction Cancers during pregnancy can be treated with chemotherapy after the first trimester but the treatment is associated with smaller placentas and an increased risk of stillbirth, fetal growth retardation and preterm delivery. We decided to assess the effect of several chemotherapeutic agents on placental development by using in vitro culture of human term cytotrophoblasts. Methods Cytotrophoblasts isolated from term placentas were cultured for 48 h and treated for 24 h with epirubicin, docetaxel, vinblastine, methotrexate, tamoxifen, 4-hydroxytamoxifen, and endoxifen. First, cell viability was assessed. Then, the effect of the treatment on trophoblast differentiation and placental angiogenesis was assessed by quantifying hCG and PlGF mRNA and protein expression. Finally, the expression of two efflux transporters, BCRP and MDR1 was investigated. Results Epirubicin only strongly decreased cell viability. Epirubicin, docetaxel, and vinblastine inhibited HCGB and PlGF expression while methotrexate, tamoxifen and its two metabolites increased it. BCRP was essentially expressed in syncytiotrophoblasts and MDR1 in undifferentiated cytotrophoblasts. Their expression was not affected by the drugs but vinblastine increased BCRP mRNA expression by 2.8-fold. Discussion The most commonly used chemotherapeutic drugs are well supported in vitro by syncytiotrophoblasts, except for epirubicin, which was very cytotoxic. Chemotherapy perturbed the expression of genes normally upregulated during placental differentiation and angiogenesis but not the expression of the drug transporters. Further studies looking at the effect of combination therapy and the transporter capacities to reject the drugs will be needed to better define the effects of chemotherapy on placental development and function.

Vascular endothelial growth factor A (VEGF-A) is one of the main growth factors involved in placental vasculogenesis and angiogenesis, but its placental expression is still ambiguous. During in vitro cultures of primary term cytotrophoblasts, VEGF could not be detected in the supernatants by enzyme-linked immunosorbent assays (ELISA). One hypothesis is that VEGF is immediately and completely bound to its soluble receptor after secretion, and cannot be recognized by the antibodies used in the commercial ELISA kits. We decided to verify this hypothesis by measuring VEGF-A expression during in vitro cultures of primary term cytotrophoblasts. Term cytotrophoblasts were cultured under 21% and 2.5% O 2 for 4 days. VEGF-A transcripts were quantified by real-time polymerase chain reaction. The proteins from cell lysates and concentrated media were separated by polyacrylamide gel electrophoresis (PAGE) under denaturing and reducing conditions, and VEGF-A immunodetected by western blotting. VEGF mRNA expression did not increase during in vitro cell differentiation under 21% O 2 , but slightly increased under 2.5% O 2 only at 24 h. VEGF-A monomer was not detected in the cell lysates and in the concentrated supernatants, while a ~ 42 KDa band corresponding to the precursor L-VEGF was detected in all the cellular extracts. Isolated term villous cytotrophoblasts produce the L-VEGF precursor but they do not secrete VEGF-A even under low-oxygen tension. The question remains about the origin of VEGF in pregnancy but also about the biological role of L-VEGF, which can represent a form of storage for rapid VEGF secretion when needed.

Objectives: To evaluate fetal growth in pregnancies complicated by placenta previa with or without placenta accreta spectrum (PAS) disorder, compared with in pregnancies with a low-lying placenta., Methods: This was a multicenter retrospective cohort study of singleton pregnancies complicated by placenta previa with or without PAS disorder, for which maternal characteristics, ultrasound-estimated fetal weight and birth weight were available. Four maternal-fetal medicine units participated in data collection of diagnosis, treatment and outcome. The control group comprised singleton pregnancies with a low-lying placenta (0.5-2 cm from the internal os). The diagnosis of PAS and depth of invasion were confirmed at delivery using both a predefined clinical grading score and histopathological examination. For comparison of pregnancy characteristics and fetal growth parameters, the study groups were matched for smoking status, ethnic origin, fetal sex and gestational age at delivery., Results: The study included 82 women with placenta previa with PAS disorder, subdivided into adherent (n = 35) and invasive (n = 47) PAS subgroups, and 146 women with placenta previa without PAS disorder. There were 64 controls with a low-lying placenta. There was no significant difference in the incidence of small-for-gestational age (SGA) (birth weight ≤ 10 th percentile) and large-for-gestational age (LGA) (birth weight ≥ 90 th  percentile) between the study groups. Median gestational age at diagnosis was significantly lower in pregnancies with placenta previa without PAS disorder than in the low-lying placenta group (P = 0.002). No significant difference was found between pregnancies complicated by placenta previa with PAS disorder and those without for any of the variables. Median estimated fetal weight percentile was significantly lower in the adherent compared with the invasive previa-PAS subgroup (P = 0.047). Actual birth weight percentile at delivery did not differ significantly between the subgroups (P = 0.804)., Conclusions: No difference was seen in fetal growth in pregnancies complicated by placenta previa with PAS disorder compared with those without and compared with those with a low-lying placenta. There was also no increased incidence of either SGA or LGA neonates in pregnancies with placenta previa and PAS disorder compared with those with placenta previa with spontaneous separation of the placenta at birth. Adverse neonatal outcome in pregnancies complicated by placenta previa and PAS disorder is linked to premature delivery and not to impaired fetal growth. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd., (Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.)

Objective: To determine whether term fetal membranes from transabdominal cerclage (TAC) patients have favorable characteristics compared with membranes from patients without TAC., Methods: A prospective study of consecutive pregnant women who had undergone TAC and were delivered by elective cesarean after 37 weeks before the onset of labor at Cliniques universitaires Saint-Luc, Brussels, between January 2015 and June 2016. Membranes were collected from two areas: overlying the cervix and located far from the cervix. Membrane thickness, 15-hydroxyprostaglandin dehydrogenase (PGDH), toll-like receptor-2 (TLR2) expression, and senescence were measured and compared between the TAC group and a control group without TAC enrolled using the same study criteria., Results: In the cervical area of the TAC group, the chorion was significantly thicker (P=0.003). PGDH and TLR2 expression were also significantly increased in the cervical area of the TAC group (P=0.021 and P=0.043, respectively). Senescence was significantly decreased in the TAC group (P=0.001)., Conclusion: A significant relationship between chorion thickening and increase in PGDH and TLR2 expression and decrease in senescence was reported in the cervical area of membranes in the TAC group. These membrane changes could prevent triggering of parturition and may account for favorable outcomes and clinical success in pregnancies with TAC., (© 2019 International Federation of Gynecology and Obstetrics.)

We report a case of vanishing gastroschisis visualized by antenatal ultrasound with a 7-year long term follow-up. Currently, the child is still dependent on daily parenteral nutrition with no signs of hepatotoxicity. To our knowledge, it's the fourth case with a long-term follow-up. Vanishing gastroschisis is a rare complication of gastroschisis. However, physicians should be aware of it because its prognosis is worse than classical gastroschisis. When a vanishing gastroschisis is visualized or suspected by antenatal ultrasound, prenatal counseling is required with explanations about the risk of short bowel syndrome, the need of parenteral nutrition and related complications (inflammatory colitis, sepsis, liver failure and organ transplant). Mortality rate was initially around 93%, and dropped to 27% after the years 2000 (versus 10% for classical gastroschisis). After birth, all children will require surgery, and sometimes autologous gastro-intestinal reconstruction. Most survivors (68%) could be taken off the TPN. Unfortunately, long-term outcomes for children with vanishing gastroschisis are still missing in current literature., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Study Question: Is 8% O2 a better percentage of atmospheric oxygen for long-term cultures of human primary term cytotrophoblasts than the conventional 21% O2 traditionally used in cell culture?, Summary Answer: Human primary term cytotrophoblasts are able to differentiate into syncytiotrophoblasts under both atmospheric oxygen levels., What Is Known Already: Cell culture is traditionally done under 21% O2, which is equal to a pO2 of ~160 mm Hg. Based on the pO2 measured after instauration of the blood circulation within the placenta, it has been proposed that cytotrophoblasts culture should be under 8% O2, which is equivalent to 60 mm Hg, and that this percentage should be considered as the physiological normoxia for cytotrophoblasts., Study, Design, Size, Duration: Cytotrophoblasts were isolated and purified from human term placentas (n > 4). Cells were cultured under 21% O2 and 8% O2 for 12 days. Several cellular parameters were assessed on Days 2, 4, 8 and 12., Participants/materials, Setting, Methods: Placentas were obtained after vaginal or elective cesarean delivery from uncomplicated pregnancies at term (n ≥ 4). Cell viability was measured by a luminescent assay based on quantitation of the ATP content of living cells. Cell fusion was assessed by quantification of syncytin and e-cadherin mRNA expression by real-time PCR and determination of the fusion index by immunofluorescent microscopy. Trophoblast differentiation was assessed by measuring the expression levels of hCGβ, inhibin α subunit (InhA) and placental growth factor (PlGF) by real-time PCR and ELISA. Finally, the effect of the two oxygen levels on apoptosis and cellular oxidative stress was also investigated by quantifying caspase 3/7 activation, superoxide dismutase 1 (SOD-1) mRNA expression and H2O2 generation., Main Results and the Role of Chance: There was no difference between 21% O2 and 8% O2 on cell viability. Cell fusion seemed to be enhanced during the first 4 days when the cells were cultured under 21% O2 compared to 8% O2. The expression level of hCGβ was equivalent in both oxygen conditions, indicating that there was no difference in trophoblast differentiation. Interestingly, InhA expression was higher under 8% O2, while PlGF expression was inhibited compared to 21% O2. This latter result indicates that 8% O2 may be more hypoxic than normoxic for in vitro culture of primary term cytotrophoblast. This is further corroborated by the fact that 21% O2 did not significantly increase caspase 3/7 activities and the oxidative stress (SOD-1 mRNA expression and H2O2 generation) in our cell cultures., Large Scale Data: Not applicable., Limitations, Reasons for Caution: The in vitro culture of cytotrophoblasts is artificial and does not reflect the in vivo situation. The cell population is nearly 100% pure, cultured as a monolayer, and the cells bath in a chemically defined culture medium deprived of any oxygen carrier. The oxygen molecules available to the cells are passively dissolved in the medium. The gas dissolution properties of the medium and the cellular consumption rate of oxygen may allow the cells to sustain a wide range of oxygen percentages from 8% to 21%., Wider Implications of the Findings: It is possible to culture human primary term cytotrophoblasts for at least 12 days. The O2 percentage of the air does not negatively affect in vitro cytotrophoblast differentiation. For in vitro culture of cytotrophoblasts, it is not necessary to lower the percentage of atmospheric oxygen to 8%., Study Funding/competing Interest(s): This work was fully supported by 'Fetus for Life' charity. The authors state that there is no conflict of interest to declare regarding the publication of this paper.

Objective: To review a single-center case series of placenta percreta and to evaluate risk factors and the impact of surgical techniques used in previous cesarean delivery., Methods: The present retrospective cohort study included pregnancies with placenta percreta managed between January 1, 2002, and March 31, 2017, at Saint Luc University Hospital, Brussels, Belgium. The data reviewed included demographics, outcomes, inter-pregnancy interval, and surgical techniques used for uterine closure in previous cesarean delivery. A cases series of non-accreta placenta previa was used as a control group., Results: There were 19 pregnancies included in the study. The most common ultrasonography signs in the study group were loss of the clear zone (14/17; 82%), placental lacunae (17/17; 100%), and subplacental hypervascularity (11/14; 79%). Median gravidity, parity, and number of previous cesarean deliveries were higher (P<0.05) and inter-pregnancy interval was longer (P<0.05) in the study group than the control group. There was no difference between the groups in the surgical techniques used for previous cesarean deliveries., Conclusion: The prenatal ultrasonography diagnosis of placenta percreta is accurate and facilitates optimal management by a specialized multidisciplinary team. Multicenter studies are required to further evaluate the impact of the surgical techniques used for prior cesarean delivery on the risks of placenta percreta in subsequent pregnancies., (© 2017 International Federation of Gynecology and Obstetrics.)

Objective: To compare perinatal outcomes following emergency cerclage between patients with singleton pregnancies with prolapsed and non-prolapsed membranes., Methods: The present retrospective cohort study included data from women who underwent physical examination-indicated emergency cerclage at between 15 and 25 weeks of pregnancy at Saint Luc University Hospital, Brussels, Belgium, between January 1, 2000, and December 31, 2014. Outcomes were compared based on the presence of prolapsed or non-prolapsed membranes. The primary outcome measures were the duration of pregnancy at delivery and the interval between cerclage and delivery. Secondary outcomes included delivery weight, fetal or neonatal death, and neonatal morbidity, including neonatal intensive care unit admission., Results: Data were included from 140 patients with cervical dilation of at least 1 cm; 85 women had non-prolapsed membranes and 55 women had prolapsed membranes. Among patients with non-prolapsed membranes, the mean duration of pregnancy at delivery was later (P<0.001), the latency between cerclage and delivery was longer (P<0.001), neonatal survival was higher (P=0.036), mean delivery weight was higher (P<0.001), the prevalence of preterm delivery was lower (P<0.001), and severe neonatal morbidity and neonatal intensive care unit admission were lower (P<0.001)., Conclusion: Having non-prolapsed membranes was associated with improved perinatal outcomes following emergency cerclage., (© 2017 International Federation of Gynecology and Obstetrics.)

Study Question: Are hypoxia-inducible factors (HIF) responsible for the potentiation of inhibin alpha subunit (INHA) gene expression in primary cultures of human term cytotrophoblasts under low-oxygen tension?, Summary Answer: Both HIF1A and endothelial PAS domain protein 1 (EPAS1) are involved in the potentiation of INHA gene upregulation in cytotrophoblasts cultured under hypoxia., What Is Known Already: During the in vitro differentiation of cytotrophoblasts into syncytiotrophoblasts under 21% O2, INHA expression increases. This expression is further increased when cells are cultured under low-oxygen tension (e.g. 2.5% O2). Moreover, in pregnancy-related diseases, such as pre-eclampsia or intrauterine growth restriction (IUGR), in which hypoxia is suspected to be responsible for the abnormal placental development, maternal serum concentration of inhibin A is elevated., Study Design, Size, Duration: Cytotrophoblasts were isolated and purified from human term placentas (n = 6). Cells were cultured under 21% O2, and allowed to differentiate for 48 h. A first group of cells was treated for 16 h under 21% O2 with dimethyloxalylglycine (DMOG) or deferoxamine (DFX), molecules that mimic hypoxia by inhibiting HIF1 proteasomal degradation. Involvement of HIF1A and EPAS1 (also known as HIF2A), two HIF isoforms expressed in trophoblasts, was shown by treating another group of cells cultured under 2.5% O2 with specific inhibitors of HIF1A and EPAS1 for 16 h. INHA mRNA expression was assessed by real-time PCR and secreted inhibin A was quantified by ELISA. The role of HIF1A and EPAS1 in INHA transcriptional regulation was further confirmed by cotransfecting primary cytotrophoblasts with a luciferase reporter plasmid containing a 3.9 kb INHA promoter and plasmids allowing overexpression of HIF1A and EPAS1., Participants/materials, Setting, Methods: Placentas were obtained after vaginal or elective cesarean delivery from uncomplicated pregnancies at term (n≥ 4). The methods used were hormone measurements in the cell supernatants by enzyme-linked immunosorbent assay, real-time quantitative PCR, western blotting, immunofluorescence microscopy and transient transfection., Main Results and the Role of Chance: HIF1 protein stabilization with DMOG and DFX increased 21% O2-induced INHA mRNA and protein upregulation (P < 0.05 versus control), while hypoxia-induced INHA upregulation was repressed by HIF1A and EPAS1 inhibitors (P < 0.05 versus control). In transfection experiments of primary term cytotrophoblasts, cloned INHA promoter transcriptional activity was increased by 2.5% O2 compared to 21% O2 (P < 0.05). Overexpression of both HIF1A and EPAS1 under 21% O2 increased cloned INHA transcriptional activity (P < 0.001 versus control)., Large Scale Data: Not applicable., Limitations, Reasons for Caution: HIF1A and EPAS1 may regulate INHA expression by binding to an hypoxia-responsive element within the promoter, but we were unable to identify such an element. Inhibition of HIF1A and EPAS1 did not completely suppress upregulation of INHA expression suggesting that other transcription factors, not identified or studied here, are involved., Wider Implications of the Findings: Our data suggest that the effect of HIF1 proteins on INHA gene promoter activity may be indirect. By demonstrating the role of HIF1A and especially EPAS1 in INHA gene upregulation under hypoxia, the results suggest that HIF1 proteins may become new therapeutic targets in the treatment of pregnancy-related diseases such as pre-eclampsia or IUGR., Study Funding/competing Interest(s): This work was fully supported by 'Fetus for Life' charity. C. Depoix was supported by a fellowship 'Fonds de Recherche Clinique' from 'Fondation Saint-Luc', Belgium. The authors declare that there is no conflict of interest regarding the publication of this paper., (© The Author 2017. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com)

During the first trimester of pregnancy, placenta formation probably occurs in a low-oxygen environment necessary to protect cytotrophoblasts from oxidative stress and to allow proper gene regulation. Transcription factors involved in gene regulation under low oxygen tension are the hypoxia-inducible factors, mainly HIF1A, EPAS1 and their dimerization partner HIF1B. Little is known about their expression during in vitro culture of cytotrophoblasts under chronic hypoxia. We assessed HIF1A and EPAS1 expression in a 4-day in vitro culture of primary term cytotrophoblasts under 21% O 2 and 2.5% O 2 . Copy numbers and relative mRNA expression were assessed by real-time quantitative polymerase chain reaction. Protein levels were quantified by immunoblot and densitometric analysis. In undifferentiated cytotrophoblasts, EPAS1 transcripts were four times more abundant than HIF1A transcripts (2.14e 7 and 5e 6 copies/μg total RNA, respectively). During cell culture, HIF1A mRNA expression increased after 24h and then decreased to stay stable. The expression was even lower when cells were grown under 2.5% O 2 . EPAS1 mRNA expression increased during cytotrophoblast differentiation. The expression was higher when cells were under 21% O 2 than when they were under 2.5% O 2 . Interestingly, HIF1A, but not EPAS1, was detected in the nuclei of undifferentiated cytotrophoblasts, and in the nuclei of cytotrophoblasts that grew under 21% O 2 . During cytotrophoblast differentiation, no variation in HIF1A protein levels was detected. To the contrary, EPAS1 protein level increased during differentiation, and oxygen tension had no effect on EPAS1 protein level. In conclusion, HIF1A and EPAS1 expression was not inhibited by chronic hypoxia during in vitro cytotrophoblast differentiation., (Copyright © 2016 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.)

The frequency of twin gestations has increased over the last few decades, mainly due to maternal age at childbearing, and the use of assisted reproductive technologies. Twins are at higher risk of aneuploidy, structural anomalies, and placental abnormalities. Some of the placental and umbilical cord abnormalities found in twin gestations are nonspecific and can be found in singleton gestations (ie, placenta previa, placental abruption, single umbilical artery, velamentous cord insertion, vasa previa, etc). However, other anomalies are unique to twin gestations, and are mainly associated with monochorionic twins-these include intraplacental anastomosis and cord entanglement. Most of these conditions can be diagnosed with ultrasound. An accurate and early diagnosis is important in the management of twin gestations. Determination of chorionicity, amnionicity, and the identification of placental anomalies are key issues for the adequate management of twin pregnancies. Pathologic placental examination after delivery can help in assessing the presence of placental and umbilical cord abnormalities, as well as providing information about chorionicity and gaining insight into the potential mechanisms of disease affecting twin gestations., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Inhibin α (Inha) gene expression is regulated, in rat granulosa cells, via a cyclic 3',5'-adenosine monophosphate (AMP)-response element (CRE) found in a region of the promoter that is homologous to the human INHA promoter. We previously found that during in vitro cytotrophoblast differentiation, human INHA gene expression was regulated by TFAP2A via association with an AP-2 site located upstream of this CRE. The aim of this study was to evaluate if the human INHA gene was also regulated by cAMP in trophoblasts, and to investigate the possible crosstalk between TFAP2 and cAMP signaling pathways in the regulation of INHA gene expression. Treatment with cAMP or forskolin increased INHA mRNA expression by 7- and 2-fold in primary cytotrophoblasts and choriocarcinoma-derived BeWo cells, respectively. Treatment with the protein kinase A inhibitor H-89 reduced forskolin-induced luciferase activity by ∼40% in BeWo cells transfected with an INHA promoter-driven luciferase reporter vector. TFAP2 overexpression increased basal luciferase activity, whereas the dominant repressor KCREB abolished it. Surprisingly, mutation of the CRE also eliminated the TFAP2-induced transcription, although TFAP2 overexpression was still able to increase forskolin-induced luciferase activity when the AP-2 binding site, but not the CRE site, was mutated. Thus, INHA gene expression is upregulated by cAMP via CRE in human trophoblasts, and TFAP2 regulates this expression by interacting with CRE., (© 2014 Wiley Periodicals, Inc.)

Timely regulated changes in oxygen partial pressure are important for placental formation. Disturbances could be responsible for pregnancy-related diseases like preeclampsia and intrauterine growth restriction. We aimed to (i) determine the effect of oxygen partial pressure on cytotrophoblast differentiation; (ii) measure mRNA expression and protein secretion from genes associated with placental angiogenesis; and (iii) determine the reversibility of these effects at different oxygen partial pressures. Term cytotrophoblasts were incubated at 21% and 2.5% O2 for 96 hr, or were switched between the two oxygen concentrations after 48 hr. Real-time PCR and enzyme-linked immunosorbent assays (ELISAs) were used to evaluate cell fusion and differentiation, measuring transcript levels for those genes involved in cell fusion and placental angiogenesis, including VEGF, PlGF, VEGFR1, sVEGFR1, sENG, INHA, and GCM1. Cytotrophoblasts underwent fusion and differentiation in 2.5% O2 . PlGF expression was inhibited while sVEGFR1 expression increased. VEGF and sENG mRNA expressions increased in 2.5% compared to 21% O2 , but no protein was detected in the cell supernatants. Finally, GCM1 mRNA expression increased during trophoblast differentiation at 21% O2 , but was inhibited at 2.5% O2 . These mRNA expression effects were reversed by returning the cells to 21% O2 . Thus, low-oxygen partial pressure does not inhibit term-cytotrophoblast cell fusion and differentiation in vitro. Lowering the oxygen partial pressure from 21% to 2.5% caused normal-term trophoblasts to reversibly modify their expression of genes associated with placental angiogenesis. This suggests that modifications observed in pregnancy diseases such as preeclampsia or growth retardation are probably due to an extrinsic effect on trophoblasts., (Copyright © 2013 Wiley Periodicals, Inc.)