Infection of pregnant women by Asian lineage strains of Zika virus (ZIKV) has been linked to brain abnormalities in their infants, yet it is uncertain when during pregnancy the human conceptus is most vulnerable to the virus. We have examined two models to study susceptibility of human placental trophoblast to ZIKV: cytotrophoblast and syncytiotrophoblast derived from placental villi at term and colonies of trophoblast differentiated from embryonic stem cells (ESC). The latter appear to be analogous to the primitive placenta formed during implantation. The cells from term placentas, which resist infection, do not express genes encoding most attachment factors implicated in ZIKV entry but do express many genes associated with antiviral defense. By contrast, the ESC-derived trophoblasts possess a wide range of attachment factors for ZIKV entry and lack components of a robust antiviral response system. These cells, particularly areas of syncytiotrophoblast within the colonies, quickly become infected, produce infectious virus and undergo lysis within 48 h after exposure to low titers (multiplicity of infection > 0.07) of an African lineage strain (MR766 Uganda: ZIKVU) considered to be benign with regards to effects on fetal development. Unexpectedly, lytic effects required significantly higher titers of the presumed more virulent FSS13025 Cambodia (ZIKVC). Our data suggest that the developing fetus might be most vulnerable to ZIKV early in the first trimester before a protective zone of mature villous trophoblast has been established. Additionally, MR766 is highly trophic toward primitive trophoblast, which may put the early conceptus of an infected mother at high risk for destruction.

Human embryonic stem cells (ESCs) readily commit to the trophoblast lineage after exposure to bone morphogenetic protein-4 (BMP-4) and two small compounds, an activin A signaling inhibitor and a FGF2 signaling inhibitor (BMP4/A83-01/PD173074; BAP treatment). During differentiation, areas emerge within the colonies with the biochemical and morphological features of syncytiotrophoblast (STB). Relatively pure fractions of mononucleated cytotrophoblast (CTB) and larger syncytial sheets displaying the expected markers of STB can be obtained by differential filtration of dispersed colonies through nylon strainers. RNA-seq analysis of these fractions has allowed them to be compared with cytotrophoblasts isolated from term placentas before and after such cells had formed syncytia. Although it is clear from extensive gene marker analysis that both ESC-and placenta-derived syncytial cells are trophoblast, each with the potential to transport a wide range of solutes and synthesize placental hormones, their transcriptome profiles are sufficiently dissimilar to suggest that the two cell types have distinct pedigrees and represent functionally different kinds of STB. We propose that the STB generated from human ESCs represents the primitive syncytium encountered in early pregnancy soon after the human trophoblast invades into the uterine wall.is encountered during at least two stages of human placental development (1-3). The first coincides with early implantation when a multinucleated syncytium forms, presumably by cell fusion events, ahead of proliferating, mononucleated, cytotrophoblast (CTB) cells originating from polar trophectoderm (3,4). This invasive syncytium emerges either during or soon after the trophoblast passes through the breached uterine epithelium and into the decidualized stromal layer beneath and appears to be responsible for hollowing out regions within the stroma to form lacunae (5), which become filled with fluid and cells from maternal blood and uterine glands and presumably provide a source of nutrients for the conceptus (3, 6). By about 12 d of gestation, soon after the blastocyst has sunk below the endometrial surface, strands of cytotrophoblast begin to form and penetrate through the primitive syncytium to form primary chorionic villi, which are subsequently invaded by extraembryonic mesoderm to form secondary and tertiary villi (villous trees) (2-4). The cytotrophoblast cells associated with the villi continue to divide and provide a progenitor cell population for the villous STB, which is the cell layer that covers the outer surface of the villi and forms the definitive interface involved in exchange of gases, nutrients, and excretory materials between the fetal placenta and maternal blood. Villous STB is also the major site for production of placental hormones. Cytotrophoblast cells at the tips of the anchoring villi proliferate and colonize the endometrium, thus expanding the placental bed and simultaneously remodeling maternal spiral arteries. The extent to which extravillous trophoblast becom...

It has been shown that adverse obstetrical outcomes such as pre-eclampsia and intrauterine growth retardation correlate with maternal infection. In this study, we investigated mechanisms involved in infection-associated abnormalities in cytotrophoblast function. Primary human first trimester cytotrophoblast cells were isolated and treated with lipopolysaccharide (LPS). Levels of the cytokines and chemokines were measured and cytotrophoblast invasion was investigated. In addition, first trimester decidual macrophages were isolated and treated with the conditioned medium from LPS-treated cytotrophoblast cells, and macrophage migration was assessed. Coculturing decidual macrophages with cytotrophoblast cells was conducted to investigate macrophage costimulatory molecule and receptor expression and intracellular cytokine production. We found that LPS exposure increased cytotrophoblast production of pro-inflammatory cytokines tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-6, and chemokines IL-8, macrophage inflammatory protein (MIP)-1alpha, and CXCL12 in a dose-dependent manner. In addition, LPS decreased cytotrophoblast invasion, and its effect was Toll-like receptor 4 (TLR4)-dependent and partly TNF-alpha-dependent. Conditioned medium from LPS-stimulated cytotrophoblast cells increased decidual macrophage migration and this effect was partly TLR4-dependent. Furthermore, coculturing decidual macrophages with LPS-exposed cytotrophoblast cells up-regulated macrophage CD80 and CD86 expression and intracellular TNF-alpha and IL-12p40 production, while down-regulating macrophage CD206 and CD209 expression and intracellular IL-10 secretion. LPS-stimulated macrophages also inhibited cytotrophoblast invasion. In conclusion, our results indicate that LPS increases the production of a subset of proinflammatory cytokines and chemokines by human first trimester cytotrophoblast cells, decreases cytotrophoblast invasion, and alters the cross talk between cytotrophoblast cells and decidual macrophages.

ProblemRecent advances in lipid research have revealed that impairments in lipid mediator signaling can be involved in the pathoetiology of a variety of diseases. We previously reported aberrant expression of autotaxin, a key enzyme for lysophosphatidic acid (LPA) production, in placentas from women with preeclampsia. The present study aimed to further explore the involvement of LPA signaling in the pathoetiology of preeclampsia.Method of studyTerm placentas were obtained from deliveries after uncomplicated pregnancy (n = 18) and those complicated by preeclampsia (n = 24). First‐trimester placental tissues were collected after elective terminations of pregnancy (n = 20). Placental expression of the six identified LPARs (LPAR1‐6) was analyzed at protein and mRNA levels.ResultsIn normal pregnancy, the mRNA expression levels of all LPARs except LPAR4 were significantly higher in term. Levels of mRNA encoding LPAR2‐5 were significantly increased in preeclampsia placentas compared with those in the normal term placentas. Using Western immunoblotting, only LPAR3 was noted to be increased at the protein level in placentas from preeclamptic pregnancies. This was validated by immunohistochemistry.ConclusionIn summary, the placental expression of LPARs, particularly LPAR3, is enhanced in preeclampsia, suggesting that disturbances in placental LPA signaling may be involved in the pathogenesis of preeclampsia.

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Electronic poster abstracts childbirth in the reference centre for fetal defects with the possibility of early cardio surgery. EP04.11 M-mode vs 2D echocardiography for measuring of fetal myocardial wall thicknessA. Sepúlveda-Martínez 1,2 , L. Objectives: M-mode and 2D modalities are both used for measuring fetal myocardial wall thicknesses. However, studies comparing the performance of both modalities are lacking. We aimed to compare the reproducibility of 2D vs M-mode for measuring myocardial wall thicknesses. Methods: A prospective study including 45 healthy fetuses from low-risk pregnancies evaluated between 18 and 41 weeks of gestation. Left and right ventricular free-wall and septal myocardial thicknesses were measured at end-diastole (d) and end-systole (s) in a transverse 4-chamber view using 2D and M-mode. All measurements were performed twice, below the level of the atrioventricular valves for three times by an experienced observer, and the mean was considered as a representative final value in each case. Intraobserver repeatability was evaluated by the concordance correlation coefficient (CCC) and both techniques were compared by t-test of the CCC. Results: Both 2D and M-mode showed a good intraobserver reproducibility for assessing myocardial wall thicknesses (table 1). Both techniques were highly correlated, with a significant better performance of M-mode in the assessment of the interventricular septum. Conclusions: Both 2D and M-mode can be used in a reproducible manner, with a slightly better performance of M-mode for assessing septal thickness. Objectives: Prospective observational study to evaluate the correlation between the feasibility of an accurate fetal heart imaging according to ISUOG practice guidelines during 2nd trimester screening and various sonographic and epidemiological aspects. Methods: Between July and December 2016, 327 singleton pregnancies were evaluated at 20th-22ndGW. Fetuses with detected heart defects were excluded. Each examination was stored, reviewed and evaluated directly offline. Each parameter of the ISUOG heart protocol was scored and characterised as adequately or non-adequately assessed by a qualified expert who performed all examinations. Adequacy of imaging for each examined parameter was correlated with maternal age and BMI, abdominal and uterus wall thickness and consistence, gestational age, placenta location and thickness, fetal spine position, orientation of heart axis, distance btw heart cross and skin, amniotic fluid btw heart and uterus wall and left or right entrance of the ultrasound beam. Results: Fetal spine position and heart axis orientation affected accurate imaging of atrial septum primum (P=.016), offsetting of AV-valves (P=.009), pulmonary veins return (P=.04) and myocardiac / endocardiac tissue (P=.007) significantly. LVOT imaging was significantly affected only by distance btw heart cross and skin (P=.004). RVOT imaging was significantly detrimentally influenced by gestational age (P=.007), distance btw heart cross and skin (P<.001) and US b...