The excitement and controversy surrounding the potential role of human embryonic stem (ES)(1,2) cells in transplantation therapy have often overshadowed their potentially more important use as a basic research tool for understanding the development and function of human tissues. Human ES cells can proliferate without a known limit and can form advanced derivatives of all three embryonic germ layers. What is less widely appreciated is that human ES cells can also form the extra-embryonic tissues that differentiate from the embryo before gastrulation. The use of human ES cells to derive early human trophoblast is particularly valuable, because it is difficult to obtain from other sources and is significantly different from mouse trophoblast. Here we show that bone morphogenetic protein 4 (BMP4), a member of the transforming growth factor-beta (TGF-beta) superfamily, induces the differentiation of human ES cells to trophoblast. DNA microarray, RT-PCR, and immunoassay analyses demonstrate that the differentiated cells express a range of trophoblast markers and secrete placental hormones. When plated at low density, the BMP4-treated cells form syncytia that express chorionic gonadotrophin (CG). These results underscore fundamental differences between human and mouse ES cells, which differentiate poorly, if at all, to trophoblast(3). Human ES cells thus provide a tool for studying the differentiation and function of early human trophoblast and could provide a new understanding of some of the earliest differentiation events of human postimplantation development.
During early pregnancy, trophoblast differentiation occurs in an environment of relative low oxygen tension which is essential for normal embryonic and placental development. At around 10-12 weeks' gestation, when the intervillous space opens to maternal blood, there is an increase in PO2. This increase correlates with the time of maximal trophoblast invasion into the maternal decidua, which allows extravillous trophoblast cells to access and remodel the maternal spiral arteries. Hypoxia Inducible Factor 1(HIF-1) is a transcription factor which activates gene transcription in response to varying oxygen concentration of cells. HIF-1 is a heterodimer composed of the inducible HIF-1alpha and the constitutively expressed HIF-1beta/ARNT subunits. Using villous explants, we hake demonstrated that the oxygen-regulated cents of early trophoblast differentiation are in part mediated by TGFbeta(3), an inhibitor of trophoblast differentiation, via HIF-1alpha. Pre-eclampsia is a disease of pregnancy that is characterized by shallow trophoblast invasion. Recently, we have reported that TGFbeta(3) is over-expressed in pre-eclamptic pregnancy and that its down-regulation restores invasive capability to trophoblast cells. Because TGFbeta(3) is downstream of HIF-1alpha, in the present study we investigated the expression of HIF-1alpha in pre-eclamptic placentae and age-matched controls using in situ hybridization and histochemical analyses. We found that HIF-1alpha mRNA and protein expression are abnormally elevated in pre-eclamptic placental tissue when compared to normal placental tissue, We conclude that pre-eclampsia may result from a developmental failure of oxygen to increase or of trophoblast cells to respond and/or sense an increase in oxygen. This will prevent the normal TGFbeta3 down-regulation and will lead to poor trophoblast invasion predisposing the pregnancy to pre-eclampsia. (C) 2002 IFPA and Elsevier Science Ltd.
In the human, fetal cytotrophoblastic cells play a key role in the implantation process and in placental development. With the progression of placentation, two pathways of differentiation lead to the formation of two distinct phenotypes. In the villous trophoblast (fusion phenotype), the trophblast differentiates from the fusion of mononuclear cytotrophoblastic cells into a syncytium, the syncytiotrophoblast. Bathing the maternal blood, the syncytiotrophoblast is involved in maternal-fetal exchanges and in placental endocrine functions. In the extravillous trophoblast (proliferative/invasive phenotype), the cytotrophoblastic cells proliferate and migrate into the decidua, remodeling the pregnant endometrium and its vasculature. This review summarizes our current knowledge of the key step of villous differentiation—the cell-cell fusion of the cytotrophoblastic cells—and on the invasion process of extravillous trophoblast. Experimental evidence demonstrates that the genetic differentiation/invasion programs of cytotrophoblastic cells could be modulated by their environment: oxygen, extracellular matrix, and soluble factors (cytokines, growth factors, and hormones). Cytotrophoblastic cells fusion and the functional differentiation of villous trophoblast are specifically stimulated by estradiol, glucocorticoids, and human chorionic gonadotropin (hCG) whereas progesterone is ineffective. Because these hormones are temporally secreted in large amounts and present at the fetomaternal interface, they are in good position to play a physiologic role in trophoblast differentiation. hCG may be important very early in pregnancy, when production of this glycoprotein is maximal, whereas estrogen increasingly produced by the fetoplacental unit and cortisol secreted from the fetal adrenal may be implicated in the end-stage maturation and aging of the trophoblast.
The human placenta is an invasive structure in which highly proliferative, migratory, and invasive extravillous trophoblast (EVT) cells migrate and invade the uterus and its vasculature. Using in vitro propagated normal first-trimester EVT cells and immortalized EVT cells, which share all of the phenotypic and functional characteristics of the normal EVT cells, it has been shown that migration/invasion of human EVT cells is stringently regulated by many growth factors, their binding proteins, extracellular matrix (ECM) components, and some adhesion molecules in an autocrine/paracrine manner at the fetal maternal interface in human pregnancy. Transforming growth factor (TGF-), decorin (a proteoglycan in the ECM), and melanoma cell adhesion molecule (Mel-CAM) inhibit, and insulin-like growth factor II (IGF-II), IGF-binding protein 1 (IGFBP-1), and endothelin 1 (ET-1) stimulate EVT cell migration/invasion. Inhibition of EVT cell migration by TGF- has been suggested to be due to upregulation of integrins, which make the cells more adhesive to the ECM. Its antiinvasive action is due to an upregulation of tissue inhibitor of matrix metalloprotease 1 (TIMP-1) and plasminogen activator inhibitor (PAI-1) and a downregulation of urokinase-type plasminogen activator (uPA). Molecular mechanisms of inhibition of migration/invasion of EVT cells by decorin and Mel-CAM remain to be identified. IGF-II action has been shown to be mediated by IGF type I receptors (IGF-RII) independently of IGF type I receptors (IGF-RI) and IGFBPs. This action of IGF-II appears to involve inhibitory G proteins and phosphorylation of mitogen-activated protein kinase (MAPK) (extracellular signal-regulated protein kinases 1 and 2 (ERK-1 and ERK-2)). IGFBP-1 stimulation of EVT cell migration appears to occur by binding its Arg-Gly-Asp (RGD) domain to 51 integrin, leading to phosphorylation of focal adhesion kinase (FAK) and MAPK (ERK-1 and ERK-2). These studies may improve our understanding of diseases related to abnormal placentation, viz. hypoinvasiveness in preeclampsia and hyperinvasiveness in trophoblastic neoplasms.Key words: trophoblast, migration, integrin, IGF-RII, IGFBP-1.Le placenta humain est une structure invasive dans laquelle les cellules du trophoblaste extravilleux (TEV), très prolifératives, migratrices et invasives, migrent et envahissent l'utérus et son système vasculaire. On a montré, en utilisant des cellules TEV du premier trimestre normales, propagées in vitro et des cellules TEV immortalisées, qui possèdent toutes les caractéristiques phénotypiques et fonctionnelles des cellules TEV normales, que la migration/invasion des cellules TEV humaines est rigoureusement régulée, de façon autocrine et paracrine, par de nombreux facteurs de croissance, leurs protéines de fixation, des composants de la matrice extracellulaire (MEC) et quelques molécules d'adhésion à l'interface fto-maternelle. Le facteur de croissance transformant (TGF)-, la décorine (protéoglycane de la MEC) et la Mel-CAM (molécule d'adhésion) inhibent, et le facteur de croissance II analogue à l'insuline (IGF)-II, la protéine de fixation de l'IGF (IGFBP)-1 et l'endothéline (ET)-1 stimulent la migration/invasion des cellules TEV. On a émis l'hypothèse que l'inhibition de la migration des cellules TEV par le TGF- est provoquée par la régulation positive des intégrines, qui augmente l'adhésion des cellules à la MEC. Son action anti-invahissante est due à une régulation positive des inhibiteurs de protéase, TIMP-1 et PAI-1, et à une régulation négative de l'uPa. Les mécanismes moléculaires de l'inhibition de la migration/invasion des cellules TEV par la décorine et la molécule Mel-CAM n'ont pas encore été identifiés. On a montré que l'action de l'IGF-II est véhiculée par le récepteur de type II de l'IGF (IGF-RII) indépendamment de l'IGF-RI et des IGFBP. Cette action mettrait en cause des protéines G inhibitrices et la phosphorylation de la protéine kinase activée par les mitogènes (MAPK) (ERK-1 et -2). La stimulation de la migration des cellules TEV par les IGFBP-1 se produirait en fixant son domaine RGD (arginine-glycine et acide aspartique) à
One major materno‐fetal interface in the human placenta is constituted by the syncytiotrophoblast, in contact with maternal blood of the intervillous space, which derives from differentiation and fusion of the villous cytotrophoblast (vct). In the present work, we purified vct from term placenta by depleting HLA class I‐ and class II‐positive cells. We found by RT‐PCR that both soluble intron 4‐retaining HLA‐G1 (sHLA‐G1) and HLA‐G2 isoforms were transcribed in purified vct. Using different HLA‐G‐specific mAb, we demonstrated by intracellular flow cytometry, Western blotting and ELISA, that sHLA‐G1 but no soluble HLA class Ia molecule was secreted by vct. We then purified sHLA‐G1 from vct culture supernatant and found that it exhibited an unusual glycosylation pattern. Finally, we showed that such trophoblast‐derived sHLA‐G1 triggered specific apoptosis of activated CD8+ T cells. Taken together, these results demonstrated that vct did secrete functional sHLA‐G1 in primary culture and suggested that, in vivo, sHLA‐G1 might be an important immunomodulatory molecule controlling the activity of maternal immune effector CD8+ cells circulating in the blood that immerses chorionic villi.
X inactivation in female mammals is one of the best studied examples of heritable gene silencing and provides an important model for studying maintenance of patterns of gene expression during differentiation and development . The process is initiated by a cis-acting RNA, the X inactive specific transcript (Xist). Xist RNA is thought to recruit silencing complexes to the inactive X, which then serve to establish and maintain the inactive state in all subsequent cell divisions [2-4]. Most lineages undergo random X inactivation, there being an equal probability of either the maternally (Xm) or paternally (Xp) inherited X chromosome being inactivated in a given cell . In the extraembryonic trophectoderm and primitive endoderm lineages of mouse embryos, however, there is imprinted X inactivation of Xp . This process is also Xist dependent . A recent study has shown that imprinted X inactivation in trophectoderm is not maintained in embryonic ectoderm development (eed) mutant mice . Here we show that Eed and a second Polycomb group protein, Enx1, are directly localized to the inactive X chromosome in XX trophoblast stem (TS) cells. The association of Eed/Enx1 complexes is mitotically stable, suggesting a mechanism for the maintenance of imprinted X inactivation in these cells.