Placental Dysfunction in Assisted Reproductive Pregnancies: Perinatal, Neonatal and Adult Life Outcomes

Obstetric and newborn outcomes of assisted reproductive technology (ART) pregnancies are associated with significative prevalence of maternal and neonatal adverse health conditions, such as cardiovascular and metabolic diseases. These data are interpreted as anomalies in placentation involving a dysregulation of several molecular factors and pathways. It is not clear which extent of the observed placental alterations are the result of ART and which originate from infertility itself. These two aspects probably act synergically for the final obstetric risk. Data show that mechanisms of inappropriate trophoblast invasion and consequent altered vascular remodeling sustain several clinical conditions, leading to obstetric and perinatal risks often found in ART pregnancies, such as preeclampsia, fetal growth restriction and placenta previa or accreta. The roles of factors such as VEGF, GATA3, PIGF, sFLT-1, sEndoglin, EGFL7, melatonin and of ART conditions, such as short or long embryo cultures, trophectoderm biopsy, embryo cryopreservation, and supraphysiologic endometrium preparation, are discussed. Inflammatory local conditions and epigenetic influence on embryos of ART procedures are important research topics since they may have important consequences on obstetric risk. Prevention and treatment of these conditions represent new frontiers for clinicians and biologists involved in ART, and synergic actions with researchers at molecular levels are advocated

Thyroid hormone-regulated molecular mechanisms driving mouse blastocyst implantation

Embryo implantation is one of the earliest and critical events of the mammalian reproduction.1 A clinical association between implantation failure and thyroid dysfunction has been extensively reported, although the molecular mechanism governing this correlation has not been elucidated yet. TH machinery is expressed in the feto-maternal unit at the time of implantation, suggesting a locaI action of TH. We have investigated the role of TH in mouse hatching and outgrowth, since dysregulation of these events induces implantation failure, leading to infertility.2 Mouse blastocysts were cultured on a feeder-layer of primary endometrial cells or on plastic, with or without TH supplementation, and hatching and outgrowth evaluated. TH stimulation was also studied on endometrial cell cultures without blastocysts. By qRT-PCR, the expression of two proteases involved in blastocyst hatching in mice (Ispl and Isp2 3,4) and MMPS was studied. TH supplementation significantly increased the number of hatching blastocysts cultured on the endometrial cells, while had limited effect on blastocysts cultured on plastic. Ispl and Isp2 were significantly up-regulated in both blastocysts and endometrial cells, independently analyzed after an overnight co­culture with TH; a less pronounced effect was observed in both blastocysts and endometrial cells cultured alone. TH also induced a significant upregulation of several MMPs in endometrial cells, independently from the presence of the blastocysts, although the coculture induced a much higher increase. TH also significantly increased the expansion of trophectodermal cells in both culture conditions, however expansion was more pronounced in the presence of the endometrial feeder layer. TH plays a key role in the bidirectional crosstalk between the competent blastocyst and the receptive endometrium at the time of implantation. TH supplementation may improve implantation success. 1. Zhang S et al. Mol Aspects Med 2013,34;939 2. Petersen CG et al. Reprod Biomed Online 2005, 10;24 3. O'Sullivan CM et al. Mol Reprod Devel 2002, 62; 328 4. Sharma et al. BMC Dev Biol 2006, 6;6

An improved in vitro model simulating the feto-maternal interface to study developmental effects of potentially toxic compounds: The example of titanium dioxide nanoparticles

The study of developmental effect of xenobiotics in humans is limited and often relies on epidemiological data. Whether and to which extent potentially toxic compounds may cross the placental barrier, and whether adverse effects on embryo development are the consequence of direct or indirect placental-mediated action is debated. The availability of in vitro models simulating the feto-maternal interface could contribute to elucidate this issue. Here, we report the development of a novel in vitro model using murine blastocyst derived trophoblast stem cells (TSC) to mimic the placental barrier and mouse embryoid bodies (EBs) to represent the embryonic tissues. We demonstrate that this model can be used for translocation studies, as well as embryotoxicity assessment of titanium dioxide nanoparticles (TiO(2)NPs). By evaluating trans-epithelial electrical resistance, translocation of fluorescein isothiocyanate-dextran beads and expression of junctional complex proteins, we show that TSCs cultured on transwell inserts under differentiating condition form syncytia. We also show that TiO(2)NPs administered in the upper transwell compartment are able to reach the lower compartment and interfere with EB differentiation when no TSC are cultured on the insert. On the contrary, when TSC are present, NPs translocate to a lesser extent and do not affect EB development. These results indicate that the proposed in vitro model is suitable to study the correlation between translocation and toxicity of TiO(2)NPs and suggest a direct effect of the particles on EB development. We propose that this model could be exploited to study developmental effect of other xenobiotics

Thyroid hormone regulates protease expression and activation of notch signaling in implantation and embryo development

A clinical association between thyroid dysfunction and pregnancy complications has been extensively reported, however the molecular mechanisms through which TH might regulate key-events of pregnancy have not been elucidated yet. In this respect, we performed in vivo studies in MMI- induced hypothyroid pregnant mice, evaluating the effect of hypothyroidism on the number of implantation sites, developing embryos/resorptions and pups per litter, at 4.5, 10.5, 18.5 days post coitum (dpc) and at birth. We also studied the expression of major molecules involved in implantation and placentation, such as the proteases ISPs, MMPs, TIMPs and Notch pathway related genes. Our results demonstrate that hypothyroidism may have a dual effect on pregnancy, by initially influencing implantation and by regulating placental development at later stages of gestation. To further elucidate the role of TH in implantation, we performed in vitro studies by culturing 3.5 dpc blastocysts in the presence of TH, with or without endometrial cells used as feeder-layer, and studied their ability to undergo hatching and outgrowth. We observed that, in the presence of endometrial feeder cells, TH is able to anticipate blastocyst hatching by up-regulating the expression of blastocyst produced ISPs, and to enhance blastocyst outgrowth by up-regulating endometrial ISPs and MMPs. These results clearly indicate that TH is involved in the bidirectional crosstalk between the competent blastocyst and the receptive endometrium at the time of implantation

Thyroid hormone regulates protease expression and activation of notch signaling in implantation and embryo development

A clinical association between thyroid dysfunction and pregnancy complications has been extensively reported, however the molecular mechanisms through which TH might regulate key-events of pregnancy have not been elucidated yet. In this respect, we performed in vivo studies in MMI- induced hypothyroid pregnant mice, evaluating the effect of hypothyroidism on the number of implantation sites, developing embryos/resorptions and pups per litter, at 4.5, 10.5, 18.5 days post coitum (dpc) and at birth. We also studied the expression of major molecules involved in implantation and placentation, such as the proteases ISPs, MMPs, TIMPs and Notch pathway related genes. Our results demonstrate that hypothyroidism may have a dual effect on pregnancy, by initially influencing implantation and by regulating placental development at later stages of gestation. To further elucidate the role of TH in implantation, we performed in vitro studies by culturing 3.5 dpc blastocysts in the presence of TH, with or without endometrial cells used as feeder-layer, and studied their ability to undergo hatching and outgrowth. We observed that, in the presence of endometrial feeder cells, TH is able to anticipate blastocyst hatching by up-regulating the expression of blastocyst produced ISPs, and to enhance blastocyst outgrowth by up-regulating endometrial ISPs and MMPs. These results clearly indicate that TH is involved in the bidirectional crosstalk between the competent blastocyst and the receptive endometrium at the time of implantation

Treatment of pregnancies complicated by intrauterine growth restriction with nitric oxide donors increases placental expression of Epidermal Growth Factor-Like Domain 7 and improves fetal growth: A pilot study

Intrauterine growth restriction (IUGR) is a pathological condition of pregnancy with high perinatal mortality and morbidity, characterized by inadequate fetal growth associated to altered maternal hemodynamics with impaired uteroplacental blood flow and placental insufficiency. To date, iatrogenic premature delivery remains the elective therapeutic strategy. However, in recent years the possibility of a therapeutic approach with vasodilators and myorelaxants, such as nitric oxide (NO) donors, has gained interest. NO controls many endothelial cell functions, including angiogenesis and vascular permeability, by regulating the expression of angiogenic factors, such as Vascular Endothelial Growth Factor. In the present study, we investigated if treatment of pregnancies complicated by IUGR with NO donors affects the expression of Epidermal Growth Factor-Like Domain 7 (EGFL7), a secreted endothelial factor, previously demonstrated to be expressed by both endothelial and trophoblast cells and involved in proper placental development. NO donor treatment induced placental levels of EGFL7 and, in association with oral fluids, significantly improved fetal growth. Ex vivo experiments confirmed that NO donors increased expression and secretion of EGFL7 by villous explants. To specifically investigate the potential response of trophoblast cells to NO, we treated HTR8-sVneo cells with NO donors and observed induction of EGFL7 expression. Altogether, our findings indicate that NO induces endothelial and trophoblast expression of EGFL7 in the placenta and improves fetal growth, suggesting a correlation between placental levels of EGFL7 and pregnancy outcome

When the Embryo Meets the Endometrium: Identifying the Features Required for Successful Embryo Implantation

Evaluation of the optimal number of embryos, their quality, and the precise timing for transfer are critical determinants in reproductive success, although still remaining one of the main challenges in assisted reproduction technologies (ART). Indeed, the success of in vitro fertilization (IVF) treatments relies on a multitude of events and factors involving both the endometrium and the embryo. Despite concerted efforts on both fronts, the overall success rates of IVF techniques continue to range between 25% and 30%. The role of the endometrium in implantation has been recently recognized, leading to the hypothesis that both the “soil” and the “seed” play a central role in a successful pregnancy. In this respect, identification of the molecular signature of endometrial receptivity together with the selection of the best embryo for transfer become crucial in ART. Currently, efforts have been made to develop accurate, predictive, and personalized tests to identify the window of implantation and the best quality embryo. However, the value of these tests is still debated, as conflicting results are reported in the literature. The purpose of this review is to summarize and critically report the available criteria to optimize the success of embryo transfer and to better understand current limitations and potential areas for improvement