scholarly journals Protein O-GlcNAcylation Promotes Trophoblast Differentiation at Implantation

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2246
Author(s):  
Peter T. Ruane ◽  
Cheryl M. J. Tan ◽  
Daman J. Adlam ◽  
Susan J. Kimber ◽  
Daniel R. Brison ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Protein Modification ◽  
Fusion Gene ◽  
Embryo Implantation ◽  
Adult Health ◽  
Trophoblast Differentiation ◽  
Human Trophoblast ◽  
Endometrial Epithelium ◽  
Glcnac Transferase

Embryo implantation begins with blastocyst trophectoderm (TE) attachment to the endometrial epithelium, followed by the breaching of this barrier by TE-derived trophoblast. Dynamic protein modification with O-linked β-N-acetylglucosamine (O-GlcNAcylation) is mediated by O-GlcNAc transferase and O-GlcNAcase (OGA), and couples cellular metabolism to stress adaptation. O-GlcNAcylation is essential for blastocyst formation, but whether there is a role for this system at implantation remains unexplored. Here, we used OGA inhibitor thiamet g (TMG) to induce raised levels of O-GlcNAcylation in mouse blastocysts and human trophoblast cells. In an in vitro embryo implantation model, TMG promoted mouse blastocyst breaching of the endometrial epithelium. TMG reduced expression of TE transcription factors Cdx2, Gata2 and Gata3, suggesting that O-GlcNAcylation stimulated TE differentiation to invasive trophoblast. TMG upregulated transcription factors OVOL1 and GCM1, and cell fusion gene ERVFRD1, in a cell line model of syncytiotrophoblast differentiation from human TE at implantation. Therefore O-GlcNAcylation is a conserved pathway capable of driving trophoblast differentiation. TE and trophoblast are sensitive to physical, chemical and nutritive stress, which can occur as a consequence of maternal pathophysiology or during assisted reproduction, and may lead to adverse neonatal outcomes and associated adult health risks. Further investigation of how O-GlcNAcylation regulates trophoblast populations arising at implantation is required to understand how peri-implantation stress affects reproductive outcomes.

scholarly journals Characterisation of Osteopontin in an In Vitro Model of Embryo Implantation

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 432 ◽  
Author(s):  
Stéphane C Berneau ◽  
Peter T Ruane ◽  
Daniel R Brison ◽  
Susan J Kimber ◽  
Melissa Westwood ◽  
...  
Keyword(s):  
Secretory Pathway ◽  
Embryo Implantation ◽  
Giant Cells ◽  
Ishikawa Cell ◽  
Altered Expression ◽  
Ishikawa Cells ◽  
Cell Layers ◽  
Endometrial Epithelium ◽  
Vitro Model

At the onset of pregnancy, embryo implantation is initiated by interactions between the endometrial epithelium and the outer trophectoderm cells of the blastocyst. Osteopontin (OPN) is expressed in the endometrium and is implicated in attachment and signalling roles at the embryo–epithelium interface. We have characterised OPN in the human endometrial epithelial Ishikawa cell line using three different monoclonal antibodies, revealing at least nine distinct molecular weight forms and a novel secretory pathway localisation in the apical domain induced by cell organisation into a confluent epithelial layer. Mouse blastocysts co-cultured with Ishikawa cell layers served to model embryo apposition, attachment and initial invasion at implantation. Exogenous OPN attenuated initial, weak embryo attachment to Ishikawa cells but did not affect the attainment of stable attachment. Notably, exogenous OPN inhibited embryonic invasion of the underlying cell layer, and this corresponded with altered expression of transcription factors associated with differentiation from trophectoderm (Gata2) to invasive trophoblast giant cells (Hand1). These data demonstrate the complexity of endometrial OPN forms and suggest that OPN regulates embryonic invasion at implantation by signalling to the trophectoderm.

scholarly journals PPARγ/RXRα Heterodimers Are Involved in Human CGβ Synthesis and Human Trophoblast Differentiation

Endocrinology ◽  
2001 ◽  
Vol 142 (10) ◽  
pp. 4504-4514 ◽  
Author(s):  
Anne Tarrade ◽  
Kristina Schoonjans ◽  
Jean Guibourdenche ◽  
Jean Michel Bidart ◽  
Michel Vidaud ◽  
...  
Keyword(s):  
Regulatory Region ◽  
Additive Effect ◽  
Placental Lactogen ◽  
Placental Development ◽  
Trophoblast Differentiation ◽  
Human Trophoblast ◽  
Transcript Levels ◽  
Nuclear Extracts

Abstract Recent studies performed with null mice suggested a role of either RXRα or PPARγ in murine placental development. We report here that both PPARγ and RXRα are strongly expressed in human villous cytotrophoblasts and syncytiotrophoblasts. Moreover, specific ligands for RXRs or PPARγ (but not for PPARα or PPARδ) increase both human CGβ transcript levels and the secretion of human CG and its free β-subunit. When combined, these ligands have an additive effect on human CG secretion. Pan-RXR and PPARγ ligands also have an additive effect on the synthesis of other syncytiotrophoblast hormones such as human placental lactogen, human placental GH, and leptin. Therefore, in human placenta, PPARγ/RXRα heterodimers are functional units during cytotrophoblast differentiation into the syncytiotrophoblast in vitro. Elements located in the regulatory region of the human CGβ gene (β5) were found to bind RXRα and PPARγ from human cytotrophoblast nuclear extracts, suggesting that PPARγ/RXRα heterodimers directly regulate human CGβ transcription. Altogether, these data show that PPARγ/RXRα heterodimers play an important role in human placental development.

scholarly journals Production of Hyaluronan by the Trophectoderm is a Prerequisite for Mouse Blastocyst Attachment

2020 ◽  
Author(s):  
Ron Hadas ◽  
Eran Gershon ◽  
Aviad Cohen ◽  
Michal Elbaz ◽  
Shifra Ben-Dor ◽  
...  
Keyword(s):  
Embryo Implantation ◽  
Uterine Epithelium ◽  
Tamoxifen Treatment ◽  
Guide Rnas ◽  
Genes Encoding ◽  
Pregnant Mice ◽  
Endometrial Epithelium ◽  
Epithelium Cells

Embryo implantation requires execution of highly synchronized processes at the feto-maternal interface, initiated by blastocyst attachment to the endometrial epithelium. Hyaluronan is a major ECM component known to regulate adhesion-associated biological processes in various physiological settings. We hypothesized that hyaluronan may facilitate blastocyst attachment. In order to test our hypothesis, we characterized the blastocyst expression of hyaluronan synthesizing and degrading enzymes, as well as the expression of hyaluronan receptors during attachment. The functional impact of hyaluronan was challenged by the use of mouse transgenic blastocysts, in which genes encoding for hyaluronan synthesizing enzymes were deleted using lentiviral incorporation of Cas-9 endonuclease alongside specific short-guide RNAs into the embryonic trophectoderm. Embryos with transgenic trophectoderm were tested for their attachment in vitro, or assessed for implantation in vivo, upon transfer to foster dams. Deletion of the trophectoderm hyaluronan biosynthesis significantly reduced the number of blastocysts attached to human uterine epithelium cells in vitro. Reduced attachment was also observed in vivo, in pregnant mice carrying blastocysts with hyaluronan-depleted trophectoderm. In agreement, trophectoderm expression of osteopontin, was downregulated upon depletion of hyaluronan. MRI measurements revealed a decrease in uterine blood vessels permeability. Uterine expression of VEGF-A, PTGS-2 and uterine osteopontin, which constitute the immediate response to blastocyst attachment was also reduced. Furthermore, impaired implantation, associated with a decrease in hyaluronan synthesis in the mural trophectoderm, obtained upon tamoxifen treatment, has been recovered by LIF administration. These results demonstrate that estrogen-regulated hyaluronan-synthesis in the trophectoderm is indispensable for mouse blastocysts attachment to the uterine epithelium.

201. Homeobox gene DLX3 regulates forskolin induced trophoblast differentiation

2008 ◽  
Vol 20 (9) ◽  
pp. 1
Author(s):  
A. Chui ◽  
B. Kalionis ◽  
S. Brennecke ◽  
P. Murthi
Keyword(s):  
Transcription Factors ◽  
Real Time ◽  
Fetal Growth ◽  
Real Time Pcr ◽  
Fetal Growth Restriction ◽  
Homeobox Gene ◽  
Trophoblast Cell ◽  
Trophoblast Differentiation ◽  
Human Trophoblast ◽  
Β Hcg

Trophoblast cells carry out important functions required for the development of the normal placenta. Disruption of these functions is associated with significant pregnancy disorders such as fetal growth restriction and pre-eclampsia. Transcription factors regulate trophoblast functions. We are interested in one such class of transcription factors known as homeobox genes. The homeobox gene Distal-less 3 (DLX3) plays a vital role in the development of the mouse placenta (Morasso, Grinberg et al. 1999) and increased levels of DLX3 have been found in placentae affected by human fetal growth restriction (Murthi and Chui, unpubl. data). However, the function of DLX3 in the human placenta is not well established. Here, we investigated whether DLX3 regulates trophoblast differentiation using a plasmid construct to overexpress DLX3 in the human trophoblast cell line, BeWo. Real-time PCR showed a significant increase in DLX3 mRNA (3.1 ± 0.1 v. 1.0 ± 0.2 control, P < 0.05, n = 3), as well as the mRNA of two known markers of differentiation 3-β-hydroxysteroid dehydrogenase (3β-HSD) (8.1 ± 1.8 v. 1.2 ± 0.1 control, P < 0.05, n = 3) and β-human chorionic gonadotropin (β-hCG) (54.9 ± 0.9 v. 49.2 ± 1.6 control, P < 0.05, n = 3). Furthermore, forskolin mediated trophoblast differentiation was verified in BeWo cells. Following forskolin induction, real-time PCR showed a significant increase in the expression of DLX3 mRNA (12.9 ± 1.2 v. 3.8 ± 0.9 control, P < 0.05, n = 4), as well as a significant increase in the mRNA expression of the differentiation markers previously tested, 3β-HSD (28.3 ± 2.4 v. 1.0 ± 0.08 control, P < 0.05, n = 4) and β-hCG (2.3 ± 1.9 v. 30.9 ± 0.08 control, P < 0.001, n = 3). The expression of an additional differentiation marker, syncytin, was also significantly increased (4.0 ± 1.9 v. 1.0 ± 0.08 control, P < 0.05, n = 4). Thus, we have shown that DLX3 is a regulator of human trophoblast cell differentiation, and that forskolin acts through DLX3 to induce trophoblast differentiation. (1) Morasso, M. I., A. Grinberg et al. (1999).

The effect of in vitro oxygen tension and days in culture on human trophoblast differentiation

2013 ◽  
Vol 100 (3) ◽  
pp. S140
Author(s):  
E.A. Bachman ◽  
S. Senapati ◽  
M.D. Sammel ◽  
S.F. Butts ◽  
M.A. Mainigi ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Trophoblast Differentiation ◽  
Human Trophoblast

scholarly journals Annexin A1/Formyl Peptide Receptor Pathway Controls Uterine Receptivity to the Blastocyst

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1188 ◽  
Author(s):  
Cristina B. Hebeda ◽  
Silvana Sandri ◽  
Cláudia M. Benis ◽  
Marina de Paula-Silva ◽  
Rodrigo A. Loiola ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Embryo Implantation ◽  
Formyl Peptide Receptor ◽  
Annexin A1 ◽  
Uterine Receptivity ◽  
Mucin 1 ◽  
Human Trophoblast ◽  
Junction Molecules ◽  
Formyl Peptide

Embryo implantation into the uterine wall is a highly modulated, complex process. We previously demonstrated that Annexin A1 (AnxA1), which is a protein secreted by epithelial and inflammatory cells in the uterine microenvironment, controls embryo implantation in vivo. Here, we decipher the effects of recombinant AnxA1 in this phenomenon by using human trophoblast cell (BeWo) spheroids and uterine epithelial cells (Ishikawa; IK). AnxA1-treated IK cells demonstrated greater levels of spheroid adherence and upregulation of the tight junction molecules claudin-1 and zona occludens-1, as well as the glycoprotein mucin-1 (Muc-1). The latter effect of AnxA1 was not mediated through IL-6 secreted from IK cells, a known inducer of Muc-1 expression. Rather, these effects of AnxA1 involved activation of the formyl peptide receptors FPR1 and FPR2, as pharmacological blockade of FPR1 or FPR1/FPR2 abrogated such responses. The downstream actions of AnxA1 were mediated through the ERK1/2 phosphorylation pathway and F-actin polymerization in IK cells, as blockade of ERK1/2 phosphorylation reversed AnxA1-induced Muc-1 and claudin-1 expression. Moreover, FPR2 activation by AnxA1 induced vascular endothelial growth factor (VEGF) secretion by IK cells, and the supernatant of AnxA1-treated IK cells evoked angiogenesis in vitro. In conclusion, these data highlight the role of the AnxA1/FPR1/FPR2 pathway in uterine epithelial control of blastocyst implantation.

scholarly journals Critical role for transcription factor AP-2α in human trophoblast differentiation

2004 ◽  
Vol 18 (1) ◽  
pp. 99-107 ◽  
Author(s):  
You-Hong Cheng ◽  
Bruce J. Aronow ◽  
Shaikh Hossain ◽  
Bruce Trapnell ◽  
Sue Kong ◽  
...  
Keyword(s):  
Dna Microarray ◽  
Critical Role ◽  
Morphological Differentiation ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Trophoblast Differentiation ◽  
Human Trophoblast ◽  
Cytotrophoblast Cells ◽  
Microarray Analyses

To examine whether AP-2α is a critical component of the genetic program that directs human trophoblast differentiation, we used DNA microarray analyses to characterize the effects of a dominant-negative form of the AP-2 protein upon in vitro differentiating cytotrophoblast cells. Human cytotrophoblast cells (>95% pure) were cultured for 3 days in the presence of control medium or medium containing an adenovirus that expresses a dominant-negative mutant of AP-2 (Ad2.AP-2D/N) or an adenovirus lacking the AP-2 mutant gene (Ad.WT). DNA microarray analyses using Affymetrix human U95Av2 GeneChips were performed on RNA extracted from the three groups of cells immediately prior to and after 3 days of cell culture. Cells infected with Ad2.AP-2D/N or Ad2.WT underwent morphological differentiation similar to that of uninfected cells, with greater than 90% of the cells in each group fusing to form multinucleated syncytiotrophoblast cells. However, Ad2.AP-2D/N markedly inhibited the induction or repression of many genes that were regulated in the noninfected and Ad2.WT-infected cells during differentiation. Eighteen of the 25 most induced genes and 17 of the 20 most repressed genes during differentiation were AP-2 dependent, with the majority of these related to extracellular organization, cellular communication, and signal transduction. Taken together, these findings strongly suggest that AP-2 plays a critical role for both the induction and repression of genes that comprise postsyncytialization gene expression programs of trophoblast differentiation and maturation. AP-2, however, is not required for the fusion of cytotrophoblast cells to form a syncytium or the expression of syncytin.

scholarly journals Interleukin-11 Promotes Migration, But Not Proliferation, of Human Trophoblast Cells, Implying a Role in Placentation

Endocrinology ◽  
2007 ◽  
Vol 148 (11) ◽  
pp. 5566-5572 ◽  
Author(s):  
Premila Paiva ◽  
Lois A. Salamonsen ◽  
Ursula Manuelpillai ◽  
Claire Walker ◽  
Alejandro Tapia ◽  
...  
Keyword(s):  
First Trimester ◽  
Extravillous Trophoblast ◽  
Hybridoma Cells ◽  
Adult Health ◽  
Signal Transducer ◽  
Human Trophoblast ◽  
And Migration ◽  
Transcription 3

Trophoblast growth and invasion of the uterine endometrium are critical events during placentation and are tightly regulated by factors produced within the trophoblast-endometrial microenvironment. Deficiencies in placentation can result in early miscarriage or preeclampsia and intrauterine growth restriction, leading to impaired fetal health. The latter has been linked to major adult health disorders. IL-11 is essential for blastocyst implantation in mice. In humans, IL-11 and its receptor IL-11 receptor α (IL-11Rα) are maximally expressed in the decidua and chorionic villi during early pregnancy; however, the role of IL-11 in trophoblast function is unknown. Therefore, we examined whether IL-11Rα is expressed in human first trimester implantation sites, and whether IL-11 influences proliferation and migration of a human extravillous trophoblast (EVT)-hybridoma cell line and primary EVT cells, used as models for EVT. Immunoreactive IL-11Rα localized to subpopulations of interstitial and endovascular EVT cells in vivo. In EVT cells in vitro, IL-11: 1) stimulated phosphorylation of signal transducer and activator of transcription-3; 2) was without effect on EVT cell proliferation; and 3) stimulated significant migration of EVT-hybridoma cells (no endogenous IL-11), whereas in primary EVT, blocking endogenous IL-11 inhibited EVT migration by 30–40%. These data demonstrate that IL-11 stimulates human EVT migration, but not proliferation, likely via signal transducer and activator of transcription-3, indicating an important role for IL-11 in placentation.

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