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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.)

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.

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.)

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.)

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.)