Maternal-placental Interface Research Articles

Postpartum hepatitis and host immunity in pregnant women with chronic HBV infection.

In order to develop immune tolerant to the fetal, maternal immune system will have some modification comparing to the time before pregnancy. Immune tolerance starts and develops at the maternal placental interface. In innate immunity, decidual natural killer (dNK) cells, macrophages and dendritic cells play a key role in immue tolerance. In adaptive immunity, a moderate increase of number and immune inhibition function of regulatory T cells (Treg) are essential for immune tolerance. The trophoblast cells and immune cells expressing indoleamine 2,3-dioxygenase (IDO), the trophoblast cells expressing HLA-G, and Th1/Th2 shifting to Th2 dominant and Th17/Treg shifting to Treg domiant are in favor of maternal fetal immune tolerance. Steroids (estrogen and progesterone) and human chorionic gonadotropin (HCG) also participate in immune tolerance by inducing Treg cells or upregulating immunosuppressive cytokines. Most of the patients with chronic HBV infection are in the "HBV immune tolerance period" before pregnancy, and the liver disease is relatively stable during pregnancy. In chronic HBV infection women, after delivery, the relative immunosuppression in vivo is reversed, and Th1 is dominant in Th1/Th2 and Th17 is dominant in Th17/Treg balance. After delivery, the number of Treg decrease and NK cells increase in quantity and cytotoxicity in peripheral blood. Liver NK cells may cause liver inflammation through a non-antigen specific mechanism. After delivery, the number of CD8+ T cells will increase and HBV specific T cell response recovers from the disfunction in pregnancy. Under the background of postpartum inflammation, the rapid decrease of cortisol after delivery, and especially the enhancement of HBV specific T cell response induced by HBV DNA and cytokines, are the main reasons for postpartum hepatitis. HBeAg positive, especially HBeAg<700 S/CO, and HBV DNA>3-5Log10IU/ml are risk factors for postpartum hepatitis. Antiviral treatment in late pregnancy can reduce the incidence of mother to child transmission (MTCT) in chronic HBV infection women. Chronic HBV infection women have hepatitis both during pregnancy and more often in 12 weeks postpartum. It is generally agreed that postpartum hepatitis is mild symptoms and self-limited. Delaying drug withdrawal to 48 weeks can increase the seroconversion rate of HBeAg in delivery women with elevated alanine aminotransferase (ALT) in pregnancy.

Zika virus impacts extracellular vesicle composition and cellular gene expression in macaque early gestation trophoblasts.

Zika virus (ZIKV) infection at the maternal–placental interface is associated with adverse pregnancy outcomes including fetal demise and pregnancy loss. To determine how infection impacts placental trophoblasts, we utilized rhesus macaque trophoblast stem cells (TSC) that can be differentiated into early gestation syncytiotrophoblasts (ST) and extravillous trophoblasts (EVT). TSCs and STs, but not EVTs, were highly permissive to productive infection with ZIKV strain DAK AR 41524. The impact of ZIKV on the cellular transcriptome showed that infection of TSCs and STs increased expression of immune related genes, including those involved in type I and type III interferon responses. ZIKV exposure altered extracellular vesicle (EV) mRNA, miRNA and protein cargo, including ZIKV proteins, regardless of productive infection. These findings suggest that early gestation macaque TSCs and STs are permissive to ZIKV infection, and that EV analysis may provide a foundation for identifying non-invasive biomarkers of placental infection in a highly translational model.

937 Adrenomedullin regulates trophoblast invasion at the uterine-placental interface and is critical for successful pregnancy

Background: Adrenomedullin (ADM) is a vasoactive peptide hormone found in adrenal medulla, central nervous system, immune cells, and muscle, and is expressed in the uterus and placenta. ADM plays important roles in angiogenesis, vasodilation, and inflammation. Altered levels of ADM have been implicated in adverse pregnancy outcomes like fetal growth restriction and pre-eclampsia. Animal models can be useful tools for examining pregnancy pathophysiology. Much of what is known about ADM in pregnancy has been learned from the mouse model; however, the rat is a superior model of human placentation due to deep hemochorial invasion. Objective: We aim to explore the role of ADM at the maternal-placental interface and characterize placentation in ADM wild type and null rat pregnancy. ADM was localized within the rat placentation site by in situ hybridization. We used Crispr-Cas9-mediated genome editing to create a mutant Adm knockout rat. The mutation targeted Exon 2 and introduced a premature stop codon, resulting in a 206 bp deletion and truncated, nonfunctional protein. Germline transmission was confirmed. Heterozygous intercrosses failed to produce live null rats. Adm heterozygous pairs were bred to examine placental and fetal growth, development, and demise through gestation. Adm null placentas/fetuses were viable at gestation day (gd) 13.5, but all had demised or resorbed by gd 16.5. Growth restriction was evident by gd 13.5 in Adm null fetuses. Many Adm null conceptus demonstrated growth restriction and fetal-placental hemorrhage and infarct. On microscopic evaluation, some ADM deficient placentation sites demonstrated failed intrauterine trophoblast invasion. Adrenomedullin is expressed in trophoblast giant cells and decidual cells in early gestation. ADM deficiency disrupts trophoblast invasion at the uterine-placental interface in the rat. Loss of Adm results in abnormalities in placentation, fetal growth restriction, and fetal death by gd 16.5. Thus, Adrenomedullin is critical for successful placentation and favorable pregnancy outcomes.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Immune tolerance at the maternal-placental interface in healthy pregnancy and pre-eclampsia.

The objective of this review is to describe the immunological mechanisms which facilitate maternal tolerance at the maternal-placental interface, and to discuss how these mechanisms are disrupted in pre-eclampsia. A literature review was performed based on the analysis of papers available on PubMed. The most important and relevant studies regarding the immunological mechanisms which facilitate maternal tolerance in healthy pregnancy and pre-eclampsia are presented in this article. The maternal-placental interface is the site where the immune tolerance begins and develops. Within the innate immunity, natural killer cells, macrophages and dendritic cells play a pivotal role in tolerance through regulation of inflammation. On the other hand, within the adaptive immunity, the correct increase of regulatory T cells is crucial for ensuring immune tolerance toward placental cells. Disturbances in maternal tolerance can lead to the appearance of pregnancy complications such as pre-eclampsia, which has a considerable impact on perinatal morbidity and mortality. Our partial knowledge of immunological mechanisms involved in tolerance at the maternal-placental interface indicates that pre-eclampsia is characterized by alterations of this maternal immune tolerance, which could represent the origin of the disease.

Placental endocrine malfunction programs ovarian defects in female murine offspring

Development of the placenta must always be in advance of that of the embryo. Evidence from domestic species demonstrates that the placenta is capable of stimulating its own development through a signalling dialogue with the endometrial glands. Placental lactogens produced by the trophoblast lead to increased expression and release of uterine secretions and mitogenic growth factors, including epidermal growth factor, that have a close temporal and spatial relationship with trophoblast proliferation. Here, we review evidence that an equivalent mechanism operates in the human. The same repertoire of receptors is present on the endometrial gland cells, and the epithelial cells have long been known to adopt a hypersecretory phenotype following an implantation. Furthermore, early pregnancy hormones stimulate the secretion of glycodelin-A and osteopontin, two ‘uterine milk proteins’ that have multiple potential effects at the maternal-placental interface, from organoid cultures derived from endometrial glands. Prolactin appears to be an important stimulant, but unlike in domestic species the human trophoblast does not secrete this hormone. Instead, it is a major product of decidual cells. Hence, complications of pregnancy that have their pathophysiological roots in deficient trophoblast proliferation may be due primarily to problems of decidualisation. Ensuring the endometrium is in an optimal state pre-conceptionally should therefore be a priority for women's health. Trophoblast stemness and proliferative capacity show a sharp decline at the switch from histotrophic to haemotrophic nutrition. This may reflect the increase in oxygen concentration or loss of growth factor support. Either way, there are implications for adaptive growth of the organ.

Genetic Variants in Preeclampsia: Lessons From Studies in Latin-American Populations.

Placental vascularization is a tightly regulated physiological process in which the maternal immune system plays a fundamental role. Vascularization of the maternal-placental interface involves a wide range of mechanisms primarily orchestrated by the fetal extravillous trophoblast and maternal immune cells. In a healthy pregnancy, an immune cross-talk between the mother and fetal cells results in the secretion of immunomodulatory mediators, apoptosis of specific cells, cellular differentiation/proliferation, angiogenesis, and vasculogenesis, altogether favoring a suitable microenvironment for the developing embryo. In the context of vasculopathy underlying common pregnancy disorders, it is believed that inefficient invasion of extravillous trophoblast cells in the endometrium leads to a poor placental blood supply, which, in turn, leads to decreased secretion of angiogenic factors, hypoxia, and inflammation commonly associated with preterm delivery, intrauterine growth restriction, and preeclampsia. In this review, we will focus on studies published by Latin American research groups, providing an extensive review of the role of genetic variants from candidate genes involved in a broad spectrum of biological processes underlying the pathophysiology of preeclampsia. In addition, we will discuss how these studies contribute to fill gaps in the current understanding of preeclampsia. Finally, we discuss some trending topics from important fields associated with pregnancy vascular disorders (e.g., epigenetics, transplantation biology, and non-coding RNAs) and underscore their possible implications in the pathophysiology of preeclampsia. As a result, these efforts are expected to give an overview of the extent of scientific research produced in Latin America and encourage multicentric collaborations by highlighted regional research groups involved in preeclampsia investigation.

Trophoblast VIP deficiency entails immune homeostasis loss and adverse pregnancy outcome in mice.

Immune homeostasis maintenance throughout pregnancy is critical for normal fetal development. Trophoblast cells differentiate into an invasive phenotype and contribute to the transformation of maternal arteries and the functional shaping of decidual leukocyte populations. Insufficient trophoblast invasion, inadequate vascular remodeling, and a loss of immunologic homeostasis are associated with pregnancy complications, such as preeclampsia and intrauterine growth restriction. Vasoactive intestinal peptide (VIP) is a pleiotropic neuropeptide synthetized in trophoblasts at the maternal-placental interface. It regulates the function of trophoblast cells and their interaction with decidual leukocytes. By means of a murine model of pregnancy in normal maternal background with VIP-deficient trophoblast cells, here we demonstrate that trophoblast VIP is critical for trophoblast function: VIP gene haploinsufficiency results in lower matrix metalloproteinase 9 expression, and reduced migration and invasion capacities. A reduced number of regulatory T cells at the implantation sites along with a lower expression of proangiogenic and antiinflammatory markers were also observed. Findings detected in the implantation sites at early stages were followed by an abnormal placental structure and lower fetal weight. This effect was overcome by VIP treatment of the early pregnant mice. Our results support the relevance of trophoblast-synthesized VIP as a critical factor in vivo for trophoblast-cell function and immune homeostasis maintenance in mouse pregnancy.-Hauk, V., Vota, D., Gallino, L., Calo, G., Paparini, D., Merech, F., Ochoa, F., Zotta, E., Ramhorst, R., Waschek, J., Leirós, C. P. Trophoblast VIP deficiency entails immune homeostasis loss and adverse pregnancy outcome in mice.

Fatty acid transporter expression and regulation is impaired in placental macrovascular endothelial cells in obese women

Objective: Fetal fatty acid (FA) delivery is ultimately controlled by placental transport. Focus has been the maternal-placental interface, but regulation at the feto-placental interface is unknown.Methods: Placental macrovascular endothelial cells (EC) (n = 4/group) and trophoblasts (TB) (n = 5/group) were isolated from lean (pregravid BMI <25 kg/m2) and obese (body mass index (BMI) > 30) women. Fatty acid transporters FAT/CD36, FABPpm, FATP4, FABP 3, 4 and 5, PLIN2 and PPARα, δ, γ expression, was measured in EC and TB. Transporter response to 24 h palmitate (PA) was assessed.Results: mRNA expression of FABP3, 4, 5 and PPARγ was 2- to 3-fold reduced in EC of obese versus lean women (p < .03), but not in TB. Protein level of FABPpm was 20% lower in obese (p < .05). Palmitate (PA) up-regulated CD36, FABP3, FABP4, and PLIN2 gene expression by 3- to 4-fold in lean but not obese EC (p < .05), while PA increased FABP4 and PLIN2 in lean and obese TB, and FABP5 in lean (p < .05) EC. PA exposure up-regulated peroxisome proliferator activated receptors (PPARs) 2-fold in lean and obese EC (p < .05), but not in TB.Conclusions: In obese women, FA transporter expression is lower in placental EC, but not TB, and less sensitive to saturated FA, compared to lean women. FA transport may be regulated at the feto-placental interface.

EP16.18: Pregnancy association of abnormal Doppler indices with inherited thrombophilia

Thrombophilia reflects a hypercoagulable condition with an increased thrombogenic potential during pregnancy. The most frequent causes of inherited thrombophilia are the factor V Leiden mutation and the prothrombin gene mutation. Repetitively, it has been hypothesised that thrombophilic mutations associate an increased risk of thrombosis at the low flow maternal-placental interface resulting in placenta-mediated complications, such as miscarriage, fetal loss, pre-eclampsia, fetal growth impairment, and abruption. Our prospective study aimed to determine the impact of thrombophilic mutations on second and third trimester umbilical and uterine artery Doppler parameters. Four hundred and forty-eight pregnant women were tested for the presence of factor V Leiden mutation, prothrombin gene mutation and also for a series of minor thrombophilic mutations. Each patient included in the study has been examined by ultrasound corresponding to its gestational age with the measurement of umbilical and uterine artery velocity. The measured parameters (pulsatility index and resistivity index) were interpreted consistent with gestational age nomograms. The incidence of factor V Leiden mutation was 10.4% in the studied group and the incidence of the prothrombin gene mutation was 4.46%; 11 cases (2.4%) expressed both major mutation. In 6.6% of cases corresponding to second trimester pregnancies with factor V Leiden mutation an increased pulsatility index of umbilical artery was found and 11% of these cases had a high pulsatility index of the uterine artery. For the cases with both major thrombophilic mutations present, Doppler parameters were increased in 4 from 6 cases. Even if inherited thrombophilia is not considered a major risk factor for most adverse pregnancy outcomes, Doppler assessment of fetal status is imperative for high-risk population. In our study, the impact of thrombophilia on Doppler indices was modest and limited to major mutations.

Expression and regulation of interleukin 6 and its receptor at the maternal-conceptus interface during pregnancy in pigs

It has been well established that interleukin 6 (IL6), a pleiotropic cytokine with multiple functional roles, is widely expressed in the female reproductive tract and mediates blastocyst implantation and placental development in many species. Uterine expression of IL6 during early pregnancy has been studied in pigs, but expression and function of IL6 at the maternal-placental interface throughout pregnancy have not been determined. Thus, we examined expression of IL6 and its receptors, IL6 receptor (IL6R) and GP130, in the uterine endometrium on Days 12 and 15 of the estrous cycle, and Days 12, 15, 30, 60, 90, and 114 of pregnancy, conceptus on Days 12 and 15, and chorioallantoic tissues on Days 30, 60, 90, and 114 of pregnancy in pigs. The expression of IL6, IL6R, and GP130 mRNA in the endometrial tissues increased dramatically during mid-to late-pregnancy and decreased at term. IL6, IL6R, and GP130 mRNAs were also expressed in conceptus and chorioallantoic tissues. Expression of IL6 mRNA was mainly localized to endometrial epithelial and stromal cells and chorioallantoic tissues, while IL6R and GP130 mRNAs were localized to glandular epithelial cells during pregnancy. The expression of IL6 mRNA was decreased by estrogen and progesterone treatment, whereas increasing doses of IL1β induced the expression of IL6 mRNA, but not IL6R and GP130 mRNAs, in endometrial tissue explants. These results indicate that expression of IL6 and its receptors at the feto-maternal interface is regulated in a stage- and cell-type-specific manner during pregnancy, suggesting that IL6 and its receptor signaling system may play an important role in the maintenance of pregnancy in pigs.

Trophoblast cells inhibit neutrophil extracellular trap formation and enhance apoptosis through vasoactive intestinal peptide-mediated pathways.

Do human trophoblast cells modulate neutrophil extracellular trap (NET) formation, reactive oxygen species (ROS) synthesis and neutrophil apoptosis through mechanisms involving vasoactive intestinal peptide (VIP)? Trophoblast cells inhibited NET formation and ROS synthesis and enhanced neutrophil apoptosis through VIP-mediated pathways in a model of maternal-placental interaction. Immune homeostasis maintenance at the maternal-placental interface is mostly coordinated by trophoblast cells. Neutrophil activation and NET formation increases in pregnancies complicated by exacerbated pro-inflammatory responses. VIP has anti-inflammatory and immunosuppressant effects and is synthesized by trophoblast cells. This is a laboratory-based observational study that sampled circulating neutrophils from 50 healthy volunteers to explore their response in vitro to factors derived from human trophoblast cells. Peripheral blood neutrophils were isolated from healthy volunteers and tested in vitro with first trimester trophoblast cell line (Swan-71 and HTR8) conditioned media (CM) or with VIP. The effect of VIP and trophoblast CM on NET formation was assessed by co-localization of elastase and DNA by confocal microscopy, DNA release and elastase activity measurement. Neutrophil apoptosis was determined by flow cytometry or fluorescence microscopy. ROS formation was assessed by flow cytometry with a fluorescent probe. VIP silencing was performed by siRNA transfection. For phagocytosis of apoptotic neutrophils, autologous monocytes were sampled, and engulfment and cytokines were assessed by flow cytometry and ELISA. Trophoblast CM and 10 nM VIP promoted neutrophil deactivation by preventing phorbol myristate acetate-induced NET formation and ROS synthesis while they increased neutrophil spontaneous apoptosis and reversed the anti-apoptotic effect of lipopolysaccharide (all P < 0.05 versus control). The effects of trophoblast CM were prevented by a VIP antagonist or when VIP knocked-down trophoblast cells were used (P < 0.05 versus control). Neutrophils driven to apoptosis by trophoblast CM could be rapidly engulfed by monocytes without increasing IL-12 production. Not applicable. The mechanisms of neutrophil deactivation by trophoblast VIP are based on the results obtained with neutrophils drawn from peripheral blood of healthy individuals interacting with trophoblast cell lines in vitro. These studies were designed to investigate biological processes at the cellular and molecular level; therefore, they have the limitations of studies in vitro and it is not possible to ascertain if these mechanisms operate similarly in vivo. We tested 50 neutrophil samples from healthy volunteers that have a normal variability in their responses. Cell lines derived from human trophoblast were used, and we cannot rule out a differential behavior of trophoblast cells in contact with neutrophils in vivo. Results presented here are consistent with an active mechanism through which neutrophils in contact with trophoblast cells would be deactivated and silently cleared by decidual macrophages throughout pregnancy. They support a novel immunomodulatory role of trophoblast VIP on neutrophils at the placenta, providing new clues for pharmacological targeting of immune and trophoblast cells in pregnancy complications associated with exacerbated inflammation. This work was funded by the National Agency of Sciences and Technology (PICT 2011-0144, 2014-0657 and 2013-2177) and University of Buenos Aires (UBACyT 20020130100040BA, 20020150100161BA and 20020130100744BA). The authors declare no competing interests.

Vasoactive Intestinal Peptide modulates trophoblast-derived cell line function and interaction with phagocytic cells through autocrine pathways.

Trophoblast cells migrate and invade the decidual stroma in a tightly regulated process to maintain immune homeostasis at the maternal-placental interface during the first weeks of pregnancy. Locally synthesized factors modulate trophoblast cell function and their interaction with maternal leukocytes to promote the silent clearance of apoptotic cells. The vasoactive intestinal peptide (VIP) is a pleiotropic polypeptide with trophic and anti-inflammatory effects in murine pregnancy models. We explored the effect of VIP on two human first trimester trophoblast cell lines, particularly on their migration, invasiveness and interaction with phagocytic cells, and the signalling and regulatory pathways involved. We found that VIP enhanced trophoblast cell migration and invasion through the activation of high affinity VPAC receptors and PKA-CRE signalling pathways. VIP knocked-down trophoblast cells showed reduced migration in basal and leukemic inhibitor factor (LIF)-elicited conditions. In parallel, VIP-silenced trophoblast cells failed to induce the phagocytosis of apoptotic bodies and the expression of immunosuppressant markers by human monocytes. Our results suggest that VIP-mediated autocrine pathways regulate trophoblast cell function and contribute to immune homeostasis maintenance at placentation and may provide new clues for therapeutic intervention in pregnancies complicated by defective deep placentation.

Trophoblast cells primed with vasoactive intestinal peptide enhance monocyte migration and apoptotic cell clearance through αvβ3 integrin portal formation in a model of maternal-placental interaction.

Is apoptotic cell phagocytosis by monocytes modulated by pathways elicited by vasoactive intestinal peptide (VIP) action on trophoblast? Targeting trophoblast cells with VIP induces monocyte migration, polarization to anti-inflammatory phenotypes and apoptotic trophoblast cell clearance which involves increased αvβ3 integrin expression on phagocytic cells and binding to thrombospondin 1. Monocytes recruited to the maternal-placental interface interact with trophoblast cells and differentiate to alternatively activated macrophages involved in the silent clearance of apoptotic cells. Vasoactive intestinal peptide (VIP) is an immunomodulatory polypeptide synthesized at the human placenta that can target both trophoblast cells and monocytes/macrophages. Integrin αvβ3 and thrombospondin 1 are involved in the formation of a phagocytic portal for the immunosuppressant clearance of apoptotic cells. This is a laboratory-based study studying monocytes isolated from peripheral blood of healthy women (n = 33) and their interaction in vitro with first trimester trophoblast cell lines. Peripheral blood monocytes were isolated from healthy volunteers by Percoll gradient and tested in co-culture settings with first trimester trophoblast cell lines (Swan 71 and HTR8) or with trophoblast cell conditioned media obtained in the presence or absence of 10 or 100 nM VIP. The effect of VIP-conditioned media on monocyte migration was assessed through transwell systems and monocyte/macrophage phenotype was determined by flow cytometry. Phagocytosis of apoptotic cells and the mechanisms involved in phagocytic portal formation were assessed by flow cytometry, confocal microscopy, immunological blockade and RT-PCR. Exposing cells to 100 nM VIP increased the migration of monocytes toward trophoblast cell conditioned media (VIP conditioned medium) (P < 0.05 versus conditioned media from cells not exposed to VIP) and contributed to the monocytes acquiring an anti-inflammatory profile with increased CD39 and IL-10 expression (P < 0.05). Phagocytosis of apoptotic trophoblast cells by monocytes and monocyte-differentiated macrophages was increased by VIP conditioned medium (P < 0.05 versus media conditioned in the absence of VIP or direct addition of 100 nM VIP). The boosting effect of VIP conditioned medium on phagocytosis involved increased expression and re-localization of αvβ3 integrin on phagocytic cells along with enhanced expression of thrombospondin 1 on trophoblast cells. The conclusions are based on in vitro experiments with monocytes drawn from peripheral blood of healthy individuals and trophoblast cell lines and we were unable to ascertain that these mechanisms operate similarly in vivo. We cannot rule out a differential behavior of either trophoblast cells targeted in vivo with VIP, or primary cultures of first trimester trophoblast cells assayed in vitro. The results presented provide new clues for immune and trophoblast cell pharmacological targeting in pregnancy complications of immunopathologic nature. This work was funded by the National Agency of Sciences and Technology ANPCyT (PICT 2011-0144), National Research Council CONICET (PIP 602/2012) and University of Buenos Aires (UBACyT 20020130100040BA) to C.P.L. The authors have no conflicts of interest to disclose.

Abstract 129: A Biopolymer-Stabilized VEGF Chimera Reverses Angiogenic Imbalance in vitro in the Reduced Uterine Perfusion Pressure Model of Preeclampsia

Preeclampsia (PE) is a hypertensive disorder that complicates approximately 5-8% of all pregnancies in the U.S. The unknown etiology of PE develops from molecular dysfunction at the maternal-placental interface, where inflammatory, oxidative stress, and anti-angiogenic pathways are initiated. The resulting hypoxic in utero environment contributes to placental ischemia from which the anti-angiogenic factor soluble fms-like tyrosine kinase-1 (sFlt-1) is released. A truncated isoform, sFlt-1 consists solely of the extracellular domain of the vascular endothelial growth factor (VEGF) receptor 1, and hence lacks both catalytic and regulatory activities. However, sFlt-1 retains a high affinity for VEGF and sequesters this ligand from binding with fully functional VEGF receptors, thus prevents the activation of angiogenic remodeling pathways. The rodent model of PE, reduced uterine perfusion pressure (RUPP), exhibits the sFlt-1 pathophysiology observed in human PE. In an effort to counteract excessive sFlt-1 production and restore angiogenic balance, we have constructed a VEGF chimera fused to an Elastin-like Polypeptide (ELP) carrier with increased in vivo half-life and stability which retains full VEGF signaling activity. When human umbilical vein vascular endothelial cells (HUVECs) are exposed to serum from normal pregnant or RUPP-treated rats, tube formation on extracellular matrix is inhibited 31% (± 2%) by the RUPP serum. This inhibition is reversed when ELP-VEGF, but not ELP control, is added to the culture medium ( p = 0.0007, one-way ANOVA with Bonferroni multiple comparison), suggesting that ELP-VEGF counteracts the anti-angiogenic factors present in RUPP serum. We also characterized the pharmacokinetics, biodistribution, and placental transfer of ELP-VEGF in the pregnant Sprague Dawley rat. These studies indicate that ELP-delivered VEGF has potential for counteracting the circulating anti-angiogenic factors in maternal plasma.

VIP boosts regulatory T cell induction by trophoblast cells in an in vitro model of trophoblast-maternal leukocyte interaction.

Inducible regulatory T cells (Tregs) exert a timely and efficient immunosuppressive action at the critical peri-implantation stage essential for maternal tolerance to the conceptus. Vasoactive intestinal peptide (VIP) promotes anti-inflammatory and tolerogenic profiles through binding to VIP receptors on immune cells. We evaluated whether VIP produced by trophoblast cells induces Tregs during the early interaction of maternal leukocytes with trophoblast cells, thus contributing to maternal tolerance. We used an in vitro model of maternal leukocyte-trophoblast cell interaction represented by cocultures of fertile women's PBMCs with a human trophoblast cell line (Swan-71) and evaluated the effect of VIP added exogenously and of the endogenous polypeptide. VIP increased the frequency of CD4(+)CD25(+)FoxP3(+) cells after coculture, and these cells were able to suppress the maternal alloresponse. VIP also increased the frequency of CD4(+)IL10(+) and CD4(+)TGFβ(+) cells, but it did not modulate IFN-γ or IL-17 production. Swan-71 secreted VIP, and their coculture with maternal PBMCs significantly increased the frequency of Tregs. This effect was even more pronounced if the trophoblast cells had been pretreated with VIP. In both situations, the VIP antagonist prevented the increase in the frequency of CD4(+)Foxp3(+) cells, reflecting a specific effect of the polypeptide after the interaction with Swan-71 cells. Finally, the increase in CD4(+)CD25(+)FoxP3(+) frequency was prevented by an anti-TGF-β Ab and a VIP antagonist. These results suggest that VIP could have an active role in the immunoregulatory processes operating in the maternal-placental interface by contributing to the induction of Tregs through a mechanism involving TGF-β1.

Acetylcholine contributes to control the physiological inflammatory response during the peri-implantation period.

Maternal antigen-presenting cells attracted to the pregnant uterus interact with trophoblast cells and modulate their functional profile to favour immunosuppressant responses. Non-neuronal cholinergic system is expressed in human cytotrophoblast cells and in immune cells with homeostatic regulatory functions. The aim of this work was to evaluate whether non-neuronal acetylcholine conditions maternal monocyte and DC migration and activation profiles. We used an in vitro model resembling maternal-placental interface represented by the co-culture of human trophoblast cells (Swan-71 cell line) and monocytes or DC. When cytotrophoblast cells were treated with neostigmine (Neo) to concentrate endogenous acetylcholine levels, monocyte migration was increased. In parallel, high levels of IL-10 and decreased levels of TNF-α were observed upon interaction of maternal monocytes with trophoblast cells. This effect was synergized by Neo and was prevented by atropine, a muscarinic acetylcholine receptor antagonist. Similarly, trophoblast cells increased the migration of DC independently of Neo treatment; however, enhanced IL-10 and MCP-1 synthesis in trophoblast-DC co-cultures with no changes in TNF-α and IL-6 was observed. In fact, there were no changes in HLA-DR, CD86 or CD83 expression. Finally, trophoblast cells treated with Neo increased the expression of two antigen-presenting cells attracting chemokines, MCP-1, MIP-1α and RANTES through muscarinic receptors, and it was prevented by atropine. Our present results support a novel role of acetylcholine synthesized by trophoblast cells to modulate antigen-presenting cell migration and activation favouring an immunosuppressant profile that contributes to immune homeostasis maintenance at the maternal-foetal interface.

Human amniotic epithelial cells: Proliferation and apoptosis during their hepatic differentiation

Human amniotic epithelial cells: Proliferation and apoptosis during their hepatic differentiation

Relevance of endogenous VIP at the early maternal-placental interface: Functional profile of trophoblast giant cells from normal and VIP-deficient mice

Relevance of endogenous VIP at the early maternal-placental interface: Functional profile of trophoblast giant cells from normal and VIP-deficient mice

Immune homeostasis maintenance from decidualization to implantation: macrophages, dendritic cells and Treg network

Immune homeostasis maintenance from decidualization to implantation: macrophages, dendritic cells and Treg network

Differential migration and activation profile of monocytes after trophoblast interaction.

Macrophages at the maternal-placental interface coordinate opposite demands under the control of trophoblast cells such as the response against pathogens on one hand, and apoptotic cell clearance and wound healing with the production of suppressor cytokines. Here, we investigated whether trophoblast cells induce maternal monocyte activation towards an alternative activated macrophage profile and whether bacterial or viral stimuli modulate their migratory properties. We used an in vitro model of the maternal-placental interface represented by co-cultures of CD14+ cells isolated from fertile women with first trimester trophoblast cell line (Swan-71 cells) in the presence or absence of pathogen associated molecular pattern (PAMP) stimuli lipopolysaccharide (LPS), peptidoglycan (PGN) or poly [I:C]). Maternal CD14+ cells showed increased CD16 and CD39 expression, both markers associated to an alternative activation profile, with no changes in CD80 expression after trophoblast cell interaction. These changes were accompanied by increased IL-10 and decreased IL-12 production by CD14+ cells. After stimulation with LPS, PGN or poly [I:C], monocytes co-cultured with trophoblast cells had lower production of TNF-α and IL-1β compared with non co-cultured monocytes. Interestingly, monocyte migration towards trophoblast cells was prevented in the presence of LPS or PGN but not after 24h of stimulation with poly [I:C]. LPS or PGN also decreased CCR5, CXCL-8 and CCL5 expression. Finally, trophoblast cells co-cultured with monocytes in the presence of pathological stimuli failed to increase chemokine expression, indicating a bidirectional effect. In conclusion, trophoblast might ‘instruct’ maternal monocytes to express an alternative activation profile and restrain their early recruitment under pathological threats as one of the first strategies to avoid potential tissue damage at the maternal-placental interface.