scholarly journals Direct Involvement of HERV-W Env Glycoprotein in Human Trophoblast Cell Fusion and Differentiation

2003 ◽  
Vol 23 (10) ◽  
pp. 3566-3574 ◽  
Author(s):  
Jean-Louis Frendo ◽  
Delphine Olivier ◽  
Valérie Cheynet ◽  
Jean-Luc Blond ◽  
Olivier Bouton ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cell Fusion ◽  
Primary Cultures ◽  
Placental Tissue ◽  
Trophoblast Cell ◽  
Direct Role ◽  
Human Trophoblast ◽  
Type D ◽  
Env Protein

ABSTRACT We recently demonstrated that the product of the HERV-W env gene, a retroviral envelope protein also dubbed syncytin, is a highly fusogenic membrane glycoprotein inducing the formation of syncytia on interaction with the type D mammalian retrovirus receptor. In addition, the detection of HERV-W Env protein (Env-W) expression in placental tissue sections led us to propose a role for this fusogenic glycoprotein in placenta formation. To evaluate this hypothesis, we analyzed the involvement of Env-W in the differentiation of primary cultures of human villous cytotrophoblasts that spontaneously differentiate by cell fusion into syncytiotrophoblasts in vitro. First, we observed that HERV-W env mRNA and glycoprotein expression are colinear with primary cytotrophoblast differentiation and with expression of human chorionic gonadotropin (hCG), a marker of syncytiotrophoblast formation. Second, we observed that in vitro stimulation of trophoblast cell fusion and differentiation by cyclic AMP is also associated with a concomitant increase in HERV-W env and hCG mRNA and protein expression. Finally, by using specific antisense oligonucleotides, we demonstrated that inhibition of Env-W protein expression leads to a decrease of trophoblast fusion and differentiation, with the secretion of hCG in culture medium of antisense oligonucleotide-treated cells being decreased by fivefold. Taken together, these results strongly support a direct role for Env-W in human trophoblast cell fusion and differentiation.

scholarly journals Trophoblast stem cells - methods of isolation, histological and cellular characteristic, and their possible applications in human and animal models

2020 ◽  
Vol 8 (3) ◽  
pp. 95-100
Author(s):  
Rafał Sibiak ◽  
Michał Jaworski ◽  
Saoirse Barrett ◽  
Rut Bryl ◽  
Paweł Gutaj ◽  
...  
Keyword(s):  
Stem Cells ◽  
Marker Gene ◽  
Experimental Studies ◽  
Placental Tissue ◽  
Trophoblast Cell ◽  
Human Trophoblast ◽  
Trophoblast Stem Cells ◽  
Transcription Profiles ◽  
Trophoblast Stem

AbstractThe placenta is a part of feto-maternal unit that develops from the maternal decidua basalis and fetal-derived trophoblast cells. The regulation of its early development is extremely intricate, albeit the elusive trophoblast stem cells (TSCs) are thought to give rise to the fetal part of the placenta. TSCs may be isolated in both animal and human models. In detail, TSCs can be efficiently obtained from the early conceptus tissues – blastocysts or early placental tissue. The isolation of murine TSCs pave the way for analyses of human trophoblast cell lineages. Both human and animal stem cells retain similar characteristic properties – the ability for unrestricted self-renewal and differentiation into all trophoblast cell lines. Nevertheless, there are some essential differences across the various species which are especially pronounced when pertaining to their distinct optimal cell culture requirements. Moreover, there are several crucial discrepancies in the stemness marker gene transcription profiles between human and murine TSCs models. In vitro TSC models can be adapted to the elucidation of the pathophysiology of various reproductive complications. For instance, their properties may illustrate the conditions observed during the implantation or simulate the state of abnormal placentation. Observations gained from the experimental studies could potentially explain the cause of some cases of infertility, preeclampsia, and fetal growth abnormalities.Running title: Update on the trophoblast stem cells

scholarly journals Overexpression of Copper Zinc Superoxide Dismutase Impairs Human Trophoblast Cell Fusion and Differentiation

Endocrinology ◽  
2001 ◽  
Vol 142 (8) ◽  
pp. 3638-3648 ◽  
Author(s):  
Jean-Louis Frendo ◽  
Patrice Thérond ◽  
Terry Bird ◽  
Nathalie Massin ◽  
Francoise Muller ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Cell Fusion ◽  
Trophoblast Cell ◽  
Human Trophoblast ◽  
Copper Zinc Superoxide Dismutase ◽  
Zinc Superoxide Dismutase ◽  
Copper Zinc

scholarly journals Activation of Peroxisome Proliferator-Activated Receptor Gamma by Human Cytomegalovirus for De Novo Replication Impairs Migration and Invasiveness of Cytotrophoblasts from Early Placentas

2009 ◽  
Vol 84 (6) ◽  
pp. 2946-2954 ◽  
Author(s):  
Benjamin Rauwel ◽  
Bernard Mariamé ◽  
Hélène Martin ◽  
Ronni Nielsen ◽  
Sophie Allart ◽  
...  
Keyword(s):  
Human Cytomegalovirus ◽  
De Novo ◽  
Present Report ◽  
Primary Cultures ◽  
First Trimester ◽  
Peroxisome Proliferator ◽  
Mobility Shift ◽  
Peroxisome Proliferator Activated Receptor ◽  
Human Trophoblast

ABSTRACT Human cytomegalovirus (HCMV) contributes to pathogenic processes in immunosuppressed individuals, in fetuses, and in neonates. In the present report, by using reporter gene activation assays and confocal microscopy in the presence of a specific antagonist, we show for the first time that HCMV infection induces peroxisome proliferator-activated receptor gamma (PPARγ) transcriptional activity in infected cells. We demonstrate that the PPARγ antagonist dramatically impairs virus production and that the major immediate-early promoter contains PPAR response elements able to bind PPARγ, as assessed by electrophoretic mobility shift and chromatin immunoprecipitation assays. Due to the key role of PPARγ in placentation and its specific trophoblast expression within the human placenta, we then provided evidence that by activating PPARγ human cytomegalovirus dramatically impaired early human trophoblast migration and invasiveness, as assessed by using well-established in vitro models of invasive trophoblast, i.e., primary cultures of extravillous cytotrophoblasts (EVCT) isolated from first-trimester placentas and the EVCT-derived cell line HIPEC. Our data provide new clues to explain how early infection during pregnancy could impair implantation and placentation and therefore embryonic development.

scholarly journals Involvement of mTOR and Regulation by AMPK in Early Iodine Deficiency-Induced Thyroid Microvascular Activation

Endocrinology ◽  
2016 ◽  
Vol 157 (6) ◽  
pp. 2545-2559 ◽  
Author(s):  
J. Craps ◽  
V. Joris ◽  
B. De Jongh ◽  
P. Sonveaux ◽  
S. Horman ◽  
...  
Keyword(s):  
Blood Flow ◽  
Protein Expression ◽  
Iodine Deficiency ◽  
Primary Cultures ◽  
Hypoxia Inducible Factor ◽  
Vascular Effects ◽  
Transient Nature ◽  
Vegf Pathway ◽  
Ribosomal S6 Kinase

Iodine deficiency (ID) induces TSH-independent microvascular activation in the thyroid via the reactive oxygen species/nitric oxide-hypoxia-inducible factor-1α/vascular endothelial growth factor (VEGF) pathway. We hypothesized the additional involvement of mammalian target of rapamycin (mTOR) as a positive regulator of this pathway and AMP-activated protein kinase (AMPK) as a negative feedback regulator to explain the transient nature of ID-induced microvascular changes under nonmalignant conditions. mTOR and AMPK involvement was investigated using an in vitro model (human thyrocytes in primary cultures) and 2 murine models of goitrogenesis (normal NMRI and RET-PTC mice [a papillary thyroid cancer model]). In NMRI mice, ID had no effect on the phosphorylation of ribosomal S6 kinase (p70S6K), a downstream target of mTOR. However, rapamycin inhibited ID-induced thyroid blood flow and VEGF protein expression. In the RET-PTC model, ID strongly increased the phosphorylation of p70S6K, whereas rapamycin completely inhibited the ID-induced increase in p70S6K phosphorylation, thyroid blood flow, and VEGF-A expression. In vitro, although ID increased p70S6K phosphorylation, the ID-stimulated hypoxia-inducible factor/VEGF pathway was inhibited by rapamycin. Activation of AMPK by metformin inhibited ID effects both in vivo and in vitro. In AMPK-α1 knockout mice, the ID-induced increase in thyroid blood flow and VEGF-A protein expression persisted throughout the treatment, whereas both parameters returned to control values in wild-type mice after 4 days of ID. In conclusion, mTOR is required for early ID-induced thyroid microvascular activation. AMPK negatively regulates this pathway, which may account for the transient nature of ID-induced TSH-independent vascular effects under benign conditions.

154. EXTRACELLULAR CALRETICULIN ALTERS ENDOTHELIAL CELL AND TROPHOBLAST CELL FUNCTIONS IN VITRO CONSISTENT WITH PRE-ECLAMPSIA

2010 ◽  
Vol 22 (9) ◽  
pp. 72
Author(s):  
N. M. Gude ◽  
K. E. Crawford ◽  
J. L. Stevenson ◽  
S. P. Brennecke
Keyword(s):  
Endothelial Cell ◽  
In Vitro Culture ◽  
Cell Number ◽  
Trophoblast Cell ◽  
Human Trophoblast ◽  
Migratory Activity ◽  
Cell Numbers ◽  
Cell Functions ◽  
Normotensive Pregnancy

Pre-eclampsia is a multisystem disorder of human pregnancy that involves abnormal placentation via insufficient trophoblast cell invasion of the maternal spiral arteries and widespread maternal endothelial cell dysfunction. Factors in plasma of pre-eclamptic women affect both trophoblast and endothelial cell functions during in vitro culture (1). The calcium-binding protein calreticulin is elevated in peripheral blood with pre-eclampsia compared to normotensive pregnancy (2). The aim of this study was to determine the effects of exogenous calreticulin at concentrations relevant to normotensive pregnancy (2 µg/mL) and to pre-eclampsia (5 µg/mL) on human trophoblast cell (HTR8) and microvascular endothelial cell (myometrial) numbers and migratory activity. Cell migration was measured by scratch assay; changes in cell number were measured by MTS assay (Promega). The results showed that calreticulin at 5µg/mL did not affect HTR8 cell number (control 68044+24542 cells, with calreticulin 72810 + 30673 cells, n = 3, P > 0.05) after 48 hours, but significantly inhibited migration of the cells by 48+11% compared to the control at 26 hours (n = 4, P < 0.02). Calreticulin at 5 µg/mL and under conditions that did not change cell number significantly increased migration of the myometrial endothelial cells by 39+7% (n = 4, P < 0.01) at 20 hours. Calreticulin at 5 µg/mL, however, significantly reduced endothelial cell numbers after 3–5 days (control 6213 + 1937 cells, with calreticulin 1937+728 cells, n = 6, P < 0.05). There was no significant change to the functions of either cell type with 2 µg/mL of calreticulin. In conclusion, exogenous calreticulin at a concentration consistent with that found in maternal blood with pre-eclampsia was shown to alter trophoblast and endothelial cell migratory activity and reduce endothelial cell numbers during in vitro culture. These results indicate that elevated circulating calreticulin may contribute to the cellular mechanisms that underlie the development of pre-eclampsia. (1) Harris et al, Reprod Sci, 2009, 16: 1082–90.(2) Gu et al, Molec Human Repro, 2008, 14: 309–15.

Comparative expression of hCG β-genes in human trophoblast from early and late first-trimester placentas

2012 ◽  
Vol 303 (8) ◽  
pp. E950-E958 ◽  
Author(s):  
M. Cocquebert ◽  
S. Berndt ◽  
N. Segond ◽  
J. Guibourdenche ◽  
P. Murthi ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Primary Cultures ◽  
First Trimester ◽  
Maternal Serum ◽  
Human Trophoblast ◽  
Intervillous Space ◽  
Β Hcg

Human chorionic gonadotropin (hCG) displays a major role in pregnancy initiation and progression and is involved in trophoblast differentiation and fusion. However, the site and the type of dimeric hCG production during the first trimester of pregnancy is poorly known. At that time, trophoblastic plugs present in the uterine arteries disappear, allowing unrestricted flow of maternal blood to the intervillous space. The consequence is an important modification of the trophoblast environment, including a rise of oxygen levels from about 2.5% before 10 wk of amenorrhea (WA) to ∼8% after 12 WA. Two specific β-hCG proteins that differ from three amino acids have been described: type 1 (CGB7) and type 2 (CGB3, -5, and -8). Here, we demonstrated in situ and ex vivo on placental villi and in vitro in primary cultures of human cytotrophoblasts that type 1 and 2 β-hCG RNAs and proteins were expressed by trophoblasts and that these expressions were higher before blood enters in the intervillous space (8–9 vs. 12–14 WA). hCG was immunodetected in villous mononucleated cytotrophoblasts (VCT) and syncytiotrophoblast (ST) at 8–9 WA but only in ST at 12–14 WA. Furthermore, hCG secretion was fourfold higher in VCT cultures from 8–9 WA compared with 12–14 WA. Interestingly, VCT from 8–9 WA placentas were found to exhibit more fusion features. Taken together, we showed that type 1 and type 2 β-hCG are highly expressed by VCT in the early first trimester, contributing to the high levels of hCG found in maternal serum at this term.

scholarly journals Characterization of the mTOR pathway in human normal adrenal and adrenocortical tumors

2014 ◽  
Vol 21 (4) ◽  
pp. 601-613 ◽  
Author(s):  
Maria Cristina De Martino ◽  
Richard A Feelders ◽  
Wouter W de Herder ◽  
Peter M van Koetsveld ◽  
Fadime Dogan ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cell Survival ◽  
Primary Cultures ◽  
Mtor Pathway ◽  
Mrna Levels ◽  
Cortisol Secretion ◽  
Specific Expression ◽  
Adrenocortical Tumors ◽  
Adrenocortical Carcinomas

The mTOR pathway has recently been suggested as a new potential target for therapy in adrenocortical carcinomas (ACCs). The aim of the current study is to describe the expression of the mTOR pathway in normal adrenals (NAs) and pathological adrenals and to explore whether there are correlation between the expression of these proteins and the in vitro response to sirolimus. For this purpose, the MTOR, S6K1 (RPS6KB1), and 4EBP1 (EIF4EBP1) mRNA expression were evaluated in ten NAs, ten adrenal hyperplasias (AHs), 17 adrenocortical adenomas (ACAs), and 17 ACCs by qPCR, whereas total(t)/phospho(p)-MTOR, t/p-S6K, and t/p-4EBP1 protein expression were assessed in three NAs, three AHs, six ACAs, and 20 ACCs by immunohistochemistry. The effects of sirolimus on cell survival and/or cortisol secretion in 12 human primary cultures of adrenocortical tumors (ATs) were also evaluated. In NAs and AHs, layer-specific expression of evaluated proteins was observed. S6K1 mRNA levels were lower in ACCs compared with NAs, AHs, and ACAs (P<0.01). A subset of ATs presented a moderate to high staining of the evaluated proteins. Median t-S6K1 protein expression in ACCs was lower than that in ACAs (P<0.01). Moderate to high staining of p-S6K1 and/or p-4EBP1 was observed in most ATs. A subset of ACCs not having moderate to high staining had a higher Weiss score than others (P<0.029). In primary AT cultures, sirolimus significantly reduced cell survival or cortisol secretion only in sporadic cases. In conclusion, these data suggest the presence of an activated mTOR pathway in a subset of ATs and a possible response to sirolimus only in certain ACC cases.

scholarly journals ARHGEF18/p114RhoGEF coordinates PKA/CREB signaling and actomyosin remodeling to drive trophoblast cell-cell fusion during placenta morphogenesis

2020 ◽  
Author(s):  
Robert Beal ◽  
Ana Alonso-Carriazo Fernandez ◽  
Dimitris K. Grammatopoulos ◽  
Karl Matter ◽  
Maria S. Balda
Keyword(s):  
Gene Expression ◽  
Cell Fusion ◽  
Rho Gtpases ◽  
Rho Gtpase ◽  
Trophoblast Cell ◽  
Cell Junctions ◽  
Nucleotide Exchange ◽  
Cell Cell

SUMMARYCoordination of cell-cell adhesion, actomyosin dynamics and gene expression is crucial for morphogenetic processes underlying tissue and organ development. Rho GTPases are main regulators of the cytoskeleton and adhesion. They are activated by guanine nucleotide exchange factors in a spatially and temporally controlled manner. However, the roles of these Rho GTPase activators during complex developmental processes are still poorly understood. ARHGEF18/p114RhoGEF is a tight junction-associated RhoA activator that forms complexes with myosin II, and regulates actomyosin contractility. Here we show that p114RhoGEF/ ARHGEF18 is required for mouse syncytiotrophoblast differentiation and placenta development. In vitro and in vivo experiments identify that p114RhoGEF controls expression of AKAP12, a protein regulating PKA signalling, and is required for PKA-induced actomyosin remodelling, CREB-driven gene expression of proteins required for trophoblast differentiation, and, hence, trophoblast cell-cell fusion. Our data thus indicate that p114RhoGEF links actomyosin dynamics and cell-cell junctions to PKA/CREB signalling, gene expression and cell-cell fusion.

Abstract 17136: MicroRNA-181b Inhibition Stabilises Abdominal Aortic Aneurysms by Promoting Collagen Accumulation and Elastin Deposition

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Karina Di Gregoli ◽  
Nur Najmi Mohamad Anuar ◽  
Sarah J George ◽  
Jason L Johnson
Keyword(s):  
Protein Expression ◽  
Collagen Fibre ◽  
Aortic Aneurysms ◽  
High Fat ◽  
Direct Role ◽  
Knockout Mouse Model ◽  
Elastin Degradation ◽  
Abdominal Aortic ◽  
Collagen Accumulation

Increased matrix metalloproteinase (MMP) activity, and related collagen and elastin degradation as a result of macrophage accumulation, are key characteristics of abdominal aortic aneurysm (AAA) progression and rupture. Accordingly targeting MMPs or promoting expression of the endogenous tissue inhibitor of metalloproteinases (TIMPs), may represent novel therapeutic strategies to prevent AAA progression and rupture. To closely mimic human AAA, we used an angiotensin II (AngII)-infused, high fat-fed apolipoprotein E knockout mouse model. Sudden death due to aortic dissection/rupture, was significantly increased (30%, p<0.001, n=20) in TIMP3 KO mice compared to wild-type controls, supporting a protective role for TIMP3 in AAA progression. Moreover Q-PCR revealed that microRNA (miR)-181b, a validated repressor of TIMP3 protein expression, was up-regulated 22-fold (n=6, p<0.001) in human AAA compared to non-aneurysmal sites. In vitro, miR181b inhibition increased macrophage TIMP3 protein expression 3-fold (n=4, p<0.05), highlighting miR181b inhibition as a potential approach to retard AAA progression. Indeed using a miR181b inhibitor, AAA development and inflammation was retarded in AngII-infused high fat-fed ApoE KO mice. This effect was characterised by a 56% reduction in macrophage content (n=5, p<0.05), augmented collagen content 1.9-fold (n=5, p<0.05) and collagen fibre thickness (+24%, p<0.05) in AAAs compared to scrambled-miR control animals. AAA elastin content was also increased (1.5-fold, n=5, p<0.05) in miR181b inhibitor treated mice versus controls. Collagen accumulation was TIMP-3 dependent as miR-181b inhibition did not affect collagen levels in TIMP-3 KO mice. However elastin content remained elevated in miR-181b inhibitor treated TIMP3 KO mice (2-fold, n=6, p=0.001), suggesting a TIMP-3 independent mechanism. In vitro miR181b inhibition in smooth muscle cells revealed a 2.5-fold increase in elastin production (n=3, p<0.05), supporting a direct role for miR-181b in elastin regulation. Collectively our data show that miR181b inhibition stabilises AAA through a dual beneficial effect, by promoting collagen preservation via TIMP3 modulation, and through directly increasing elastin production and deposition.

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