Different regulation of IRE1α and eIF2α pathways by oxygen and insulin in ACH-3P trophoblast model

Reproduction ◽  
2021 ◽  
Author(s):  
Veronika Tandl ◽  
Denise Hoch ◽  
Julia Bandres-Meriz ◽  
Sanela Nikodijevic ◽  
Gernot Desoye ◽  
...  
Keyword(s):  
Er Stress ◽  
Early Pregnancy ◽  
Physiological Role ◽  
Brefeldin A ◽  
First Trimester ◽  
Fold Change ◽  
Low Oxygen ◽  
Placental Development ◽  
Eif2α Phosphorylation ◽  
The Unfolded Protein Response

Endoplasmic reticulum (ER)-stress activates the unfolded protein response (UPR), which plays a (patho)physiological role in the placenta. Oxygen and hyperinsulinemia are major regulators of placental development. Thus, we hypothesized that oxygen, insulin and their interplay modulate ER-stress in early pregnancy. Using the human first trimester trophoblast cell line ACH-3P, we quantified mRNA and protein of several members of UPR by RT-qPCR and Western blotting, respectively. ER-stress induction using tunicamycin and brefeldin A resulted in increased CHOP (4.6-fold change; P ≤ 0.001), XBP1 expression (1.7- and 1.3-fold change, respectively; P ≤ 0.001 and P < 0.05) and XBP1 splicing (7.9- and 12.8-fold change, respectively; P ≤ 0.001). We subsequently analyzed the effect of oxygen (6.5%, 2.5%), insulin (0.1-10 nM) and their interaction using ANCOVA adjusted for cell passage as co-variate. Although GRP78 protein remained unaffected, low oxygen (2.5% O2) increased IRE1α phosphorylation (+52%; P < 0.05) and XBP1 splicing (1.8-fold change; P ≤ 0.001) after 24h, while eIF2α protein and CHOP expression were downregulated (–28%; P < 0.05 and –24%; P ≤ 0.001; respectively). eIF2α phosphorylation was also reduced after 48h by low oxygen (–61%; P < 0.05), but increased in the presence of insulin (+46%; P ≤ 0.01). These changes were not PERK-mediated, since PERK phosphorylation and total protein were not altered. Overall, our results suggest that IRE1α and eIF2α UPR-pathways are differentially regulated by oxygen and insulin in early pregnancy.

INFLUENCE OF GLUCOSE AND INSULIN OF PROSTACYCLIN SYNTHESIS IN CULTURED TR0PH0BLAST CELLS

1987 ◽  
Author(s):  
I Rákóczi ◽  
Gy Geró ◽  
J Demeter ◽  
I Gáti
Keyword(s):  
New England ◽  
Early Pregnancy ◽  
Horse Serum ◽  
Physiological Role ◽  
First Trimester ◽  
Serum Glucose ◽  
Trophoblast Invasion ◽  
Trophoblast Cells ◽  
Specific Radioimmunoassay ◽  
High Concentrations

It is known that placental cells can produce prostacyclin /PGI2/. At present, the physiological role of trophoblast PGI2 production can only be speculative. PGI2 produced by trophoblast may help to prevent the platelet aggregation and thrombosis in spiral arteries and it can also explain the maintance of, and blood flow through, the spiral arteries during endovascular trophoblast invasion in early pregnancy. It has been previously shown that increased glucose concentrations in the incubation fluid can inhibit the formation of PGI2 by rat aortic rings. The aim of present investigation was to study the effect of glucose and insulin on the generation of PGI? by trophoblast obtained from early pregnancy. Trophoblast tissue was obtained immediately from surgical termination of first trimester pregnancy /9 specimens/. Trophoblast was cultured using the method of Jogee et al. Cells obtained from trypsinizaticn were cultured at a density of 2×105 cells/ml in medium 199 containing 10% horse,serum. Glucose /5.5, 16.5 and 33mmol/l / and insulin /103 , 104 and 106 mU/1 / were added to culture and the effect on 6-oxo-PGF2 production over a 24-hr incubation was assessed. Control cultures were incubated without glucose and insulin. The concentration of 6-oxo-PGF2 in culture supernatans were measured by specific radioimmunoassay /3H-6-oxo-PGF1, RIA-kit, New England Nuclear, USA /.There was a significant decrease in 6-keto-PGF10c production by trophoblast cells incubated with increased glucose concentrations /16.5 and 33 mmol/1 / compared to controls / p<0.001/. In contrast, insulin in growth medium did not have any effects on the PGI production. These data suggest that high concentrations of 2 glucose inhibit PGIp production by cultured trophoblast cells. This decreased PGIp synthesis may impair the blood supply of trophoblast which could play a role in the development of congenital anomalies in pregnant women with poorly controlled diabetes mellitus in the first trimester of pregnancy.

scholarly journals Trophoblast lineage specification, differentiation and their regulation by oxygen tension

2018 ◽  
Vol 236 (1) ◽  
pp. R43-R56 ◽  
Author(s):  
Ching-Wen Chang ◽  
Anna K Wakeland ◽  
Mana M Parast
Keyword(s):  
Oxygen Tension ◽  
First Trimester ◽  
Arterial Blood Flow ◽  
Extravillous Trophoblast ◽  
Low Oxygen ◽  
Epithelial Stem Cells ◽  
Placental Development ◽  
Trophoblast Differentiation ◽  
Arterial Blood ◽  
Low Oxygen Tension

Development of the early embryo takes place under low oxygen tension. Under such conditions, the embryo implants and the trophectoderm, the outer layer of blastocyst, proliferate, forming the cytotrophoblastic shell, the early placenta. The cytotrophoblasts (CTBs) are the so-called epithelial ‘stem cells’ of the placenta, which, depending on the signals they receive, can differentiate into either extravillous trophoblast (EVT) or syncytiotrophoblast (STB). EVTs anchor the placenta to the uterine wall and remodel maternal spiral arterioles in order to provide ample blood supply to the growing fetus. STBs arise through CTB fusion, secrete hormones necessary for pregnancy maintenance and form a barrier across which nutrient and gas exchange can take place. The bulk of EVT differentiation occurs during the first trimester, before the onset of maternal arterial blood flow into the intervillous space of the placenta, and thus under low oxygen tension. These conditions affect numerous signaling pathways, including those acting through hypoxia-inducible factor, the nutrient sensor mTOR and the endoplasmic reticulum stress-induced unfolded protein response pathway. These pathways are known to be involved in placental development and disease, and specific components have even been identified as directly involved in lineage-specific trophoblast differentiation. Nevertheless, much controversy surrounds the role of hypoxia in trophoblast differentiation, particularly with EVT. This review summarizes previous studies on this topic, with the intent of integrating these results and synthesizing conclusions that resolve some of the controversy, but then also pointing to remaining areas, which require further investigation.

scholarly journals Proteome Profiling of Human Placenta Reveals Developmental Stage-Dependent Alterations in Protein Signature

2021 ◽  
Author(s):  
Sara Khorami Sarvestani ◽  
Sorour Shojaeian ◽  
Negar Vanaki ◽  
Behrouz Gharesi-Fard ◽  
Mehdi Amini ◽  
...  
Keyword(s):  
Human Placenta ◽  
First Trimester ◽  
Full Term ◽  
Chemical Stimulus ◽  
Health It ◽  
Low Oxygen ◽  
Placental Development ◽  
Liquid Chromatography Tandem Mass ◽  
Complex Organ ◽  
Protein Signature

Abstract Introduction Placenta is a complex organ that plays a significant role in the maintenance of pregnancy health. It is a dynamic organ that undergoes dramatic changes in growth and development at different stages of gestation. In the first-trimester, the conceptus develops in a low oxygen environment that favors organogenesis in the embryo and cell proliferation and angiogenesis in the placenta; later in pregnancy, higher oxygen concentration is required to support the rapid growth of the fetus. This transition, which appears unique to the human placenta, must be finely tuned through successive rounds of protein signature alterations. This study compares placental proteome in normal first-trimester (FT) and term human placentas (TP). MethodsNormal human first-trimester and term placental samples were collected and differentially expressed proteins were identified using two-dimensional liquid chromatography-tandem mass spectrometry.ResultsDespite the overall similarities, 120 proteins were differently expressed in first and term placentas. Out of these, 120 proteins, expression of 72 was up-regulated and that of 48 was down-regulated in the first when compared with the full term placentas. Twenty out of 120 differently expressed proteins were sequenced, among them seven showed increased (GRP78, PDIA3, ENOA, ECH1, PRDX4, ERP29, ECHM), eleven decreased (TRFE, ALBU, K2C1, ACTG, CSH2, PRDX2, FABP5, HBG1, FABP4, K2C8, K1C9) expression in first-trimester compared to the full-term placentas and two proteins exclusively expressed in first-trimester placentas (MESD, MYDGF). DiscussionAccording to Reactome and PANTHER softwares, these proteins were mostly involved in response to chemical stimulus and stress, regulation of biological quality, programmed cell death, hemostatic and catabolic processes, protein folding, cellular oxidant detoxification, coagulation and retina homeostasis. Elucidation of alteration in protein signature during placental development would provide researchers with a better understanding of the critical biological processes of placentogenesis and delineate proteins involved in regulation of placental function during development.

scholarly journals IRE1α-Dependent Decay of CReP/Ppp1r15b mRNA Increases Eukaryotic Initiation Factor 2α Phosphorylation and Suppresses Protein Synthesis

2015 ◽  
Vol 35 (16) ◽  
pp. 2761-2770 ◽  
Author(s):  
Jae-Seon So ◽  
Sungyun Cho ◽  
Sang-Hyun Min ◽  
Scot R. Kimball ◽  
Ann-Hwee Lee
Keyword(s):  
Protein Synthesis ◽  
Er Stress ◽  
Translational Control ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Regulatory Subunit ◽  
Dependent Manner ◽  
Eukaryotic Translation ◽  
Eif2α Phosphorylation ◽  
The Unfolded Protein Response

The unfolded protein response (UPR) regulates endoplasmic reticulum (ER) homeostasis and protects cells from ER stress. IRE1α is a central regulator of the UPR that activates the transcription factor XBP1s through an unconventional splicing mechanism using its endoribonuclease activity. IRE1α also cleaves certain mRNAs containing XBP1-like secondary structures to promote the degradation of these mRNAs, a process known as regulated IRE1α-dependent decay (RIDD). We show here that the mRNA of CReP/Ppp1r15b, a regulatory subunit of eukaryotic translation initiation factor 2α (eIF2α) phosphatase, is a RIDD substrate. eIF2α plays a central role in the integrated stress response by mediating the translational attenuation to decrease the stress level in the cell. CReP expression was markedly suppressed in XBP1-deficient mice livers due to hyperactivated IRE1α. Decreased CReP expression caused the induction of eIF2α phosphorylation and the attenuation of protein synthesis in XBP1-deficient livers. ER stress also suppressed CReP expression in an IRE1α-dependent manner, which increased eIF2α phosphorylation and consequently attenuated protein synthesis. Taken together, the results of our study reveal a novel function of IRE1α in the regulation of eIF2α phosphorylation and the translational control.

scholarly journals O-GlcNAc signaling attenuates ER stress-induced cardiomyocyte death

2009 ◽  
Vol 297 (5) ◽  
pp. H1711-H1719 ◽  
Author(s):  
Gladys A. Ngoh ◽  
Tariq Hamid ◽  
Sumanth D. Prabhu ◽  
Steven P. Jones
Keyword(s):  
Cell Death ◽  
Er Stress ◽  
Cardiac Myocyte ◽  
Brefeldin A ◽  
Unfolded Protein ◽  
Homologous Protein ◽  
Stress Markers ◽  
The Unfolded Protein Response ◽  
Glcnac Transferase ◽  
Protein Response

We previously demonstrated that the O-linked β- N-acetylglucosamine ( O-GlcNAc) posttranslational modification confers cardioprotection at least partially through mitochondrial-dependent mechanisms, but it remained unclear if O-GlcNAc signaling interfered with other mechanisms of cell death. Because ischemia/hypoxia causes endoplasmic reticulum (ER) stress, we ascertained whether O-GlcNAc signaling could attenuate ER stress-induced cell death per se. Before induction of ER stress (with tunicamycin or brefeldin A), we adenovirally overexpressed O-GlcNAc transferase (AdOGT) or pharmacologically inhibited O-GlcNAcase [via O-(2-acetamido-2-deoxy-d-glucopyranosylidene) amino- N-phenylcarbamate] to augment O-GlcNAc levels or adenovirally overexpressed O-GlcNAcase to reduce O-GlcNAc levels. AdOGT significantly ( P < 0.05) attenuated the activation of the maladaptive arm of the unfolded protein response [according to C/EBP homologous protein (CHOP) activation] and cardiomyocyte death (reflected by percent propidium iodide positivity). Moreover, pharmacological inhibition of O-GlcNAcase significantly ( P < 0.05) mitigated ER stress-induced CHOP activation and cardiac myocyte death. Interestingly, overexpression of GCA did not alter ER stress markers but exacerbated brefeldin A-induced cardiomyocyte death. We conclude that enhanced O-GlcNAc signaling represents a partially proadaptive response to reduce ER stress-induced cell death. These results provide new insights into a possible interaction between O-GlcNAc signaling and ER stress and may partially explain a mechanism of O-GlcNAc-mediated cardioprotection.

scholarly journals The endocannabinoid 2-arachidonoylglycerol promotes endoplasmic reticulum stress in placental cells

Reproduction ◽  
2020 ◽  
Vol 160 (2) ◽  
pp. 171-180 ◽  
Author(s):  
Marta Almada ◽  
Lia Costa ◽  
Bruno Fonseca ◽  
Patrícia Alves ◽  
Jorge Braga ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Cannabinoid Receptor ◽  
Initiation Factor ◽  
Parp Cleavage ◽  
Ros Generation ◽  
Placental Development ◽  
Induced Apoptosis ◽  
The Unfolded Protein Response ◽  
Er Homeostasis

Proliferation, differentiation and apoptosis of trophoblast cells are required for normal placental development. Impairment of those processes may lead to pregnancy-related diseases. Disruption of endoplasmic reticulum (ER) homeostasis has been associated with several reproductive pathologies including recurrent pregnancy loss and preeclampsia. In the unfolded protein response (UPR), specific ER-stress signalling pathways are activated to restore ER homeostasis, but if the adaptive response fails, apoptosis is triggered. Protein kinase RNA-like endoplasmic reticulum kinase (PERK), inositol-requiring enzyme 1 (IRE1) and Activating transcription factor 6 (ATF6) are central players in UPR and in ER-stress-induced apoptosis, as well as downstream transcription factors, as C/EBP homologous protein (CHOP). Our previous studies have shown that the endocannabinoid 2-arachidonoylglycerol (2-AG) modulates trophoblast cell turnover. Nevertheless, the role of ER-stress on 2-AG induced apoptosis and cannabinoid signalling in trophoblast has never been addressed. In this work, we used BeWo cells and human primary cytotrophoblasts isolated from term-placenta. The expression of ER-stress markers was analysed by qRT-PCR and Western blotting. ROS generation was assessed by fluorometric methods, while apoptosis was detected by the evaluation of caspase -3/-7 activities and Poly (ADP-ribose) polymerase (PARP) cleavage. Our findings indicate that 2-AG is able to induce ER-stress and apoptosis. Moreover, the eukaryotic initiation factor 2 (eIF2α)/CHOP pathway involved in ER-stress-induced apoptosis is triggered through a mechanism dependent on cannabinoid receptor CB2 activation. The results bring novel insights on the importance of ER-stress and cannabinoid signalling on 2-AG mechanisms of action in placenta.

scholarly journals Reference ranges of AMH in early pregnancy: the Generation R Study, a population-based prospective cohort study

2021 ◽  
Author(s):  
Ramon Dykgraaf ◽  
Joop Laven ◽  
Sarah Timmermans ◽  
Maria Adank ◽  
Sjoerd van den Berg ◽  
...  
Keyword(s):  
Cohort Study ◽  
Prospective Cohort Study ◽  
Early Pregnancy ◽  
Gestational Age ◽  
Prospective Cohort ◽  
Hospital Setting ◽  
First Trimester ◽  
Population Based ◽  
Reference Ranges ◽  
Placental Development

Objective: The objective of this study is to establish maternal reference values of AMH in a fertile multi-ethnic urban pregnant population and to evaluate the effect of gestational age. Design: The Generation R Study is an ongoing population-based prospective cohort study from early pregnancy onwards. Setting: Rotterdam, the Netherlands, out of hospital setting. Population: In 5806 women serum AMH levels were determined in early pregnancy (median 13.5 weeks; 95% range 10.5-17.2). Methods: The model-based AMH reference ranges for maternal age and gestational age were created using GAMLSS. Associations between AMH and several first trimester biomarkers were analyzed using multivariate linear regression analyses. Main outcome measures: AMH levels in early pregnancy and the association with placental biomarkers, hCG, sFLT, and PLGF. Results: A nomogram of AMH in early pregnancy was developed. Serum AMH levels showed a decline with advancing gestational age. Higher AMH levels were associated with a higher level of hCG and sFLT. This last association was predominantly mediated by hCG. AMH levels were negatively associated with PLGF levels. Conclusion: In this large study we show that AMH levels in the first trimester decrease with advancing gestational age. The association between AMH and the placental biomarkers hCG, sFLT and PLGF suggests a better placental development with a lower vascular resistance in mothers with higher AMH levels. AMH might be useful in predicting adverse pregnancy outcome due to impaired placental development. Keywords: Ovarian reserve, placental biomarker, nomogram, first trimester, human Choriogonadotrophin (hCG), soluble FMS-Like Tyrosine kinase-1 (sFLT), Placental Growth Factor (PLGF).

scholarly journals Proteome profiling of human placenta reveals developmental stage-dependent alterations in protein signature

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Sara Khorami Sarvestani ◽  
Sorour Shojaeian ◽  
Negar Vanaki ◽  
Behrouz Ghresi-Fard ◽  
Mehdi Amini ◽  
...  
Keyword(s):  
Human Placenta ◽  
First Trimester ◽  
Full Term ◽  
Chemical Stimulus ◽  
Health It ◽  
Low Oxygen ◽  
Placental Development ◽  
Liquid Chromatography Tandem Mass ◽  
Normal Human ◽  
Protein Signature

Abstract Introduction Placenta is a complex organ that plays a significant role in the maintenance of pregnancy health. It is a dynamic organ that undergoes dramatic changes in growth and development at different stages of gestation. In the first-trimester, the conceptus develops in a low oxygen environment that favors organogenesis in the embryo and cell proliferation and angiogenesis in the placenta; later in pregnancy, higher oxygen concentration is required to support the rapid growth of the fetus. This oxygen transition, which appears unique to the human placenta, must be finely tuned through successive rounds of protein signature alterations. This study compares placental proteome in normal first-trimester (FT) and term human placentas (TP). Methods Normal human first-trimester and term placental samples were collected and differentially expressed proteins were identified using two-dimensional liquid chromatography-tandem mass spectrometry. Results Despite the overall similarities, 120 proteins were differently expressed in first and term placentas. Out of these, 72 were up-regulated and 48 were down-regulated in the first when compared with the full term placentas. Twenty out of 120 differently expressed proteins were sequenced, among them seven showed increased (GRP78, PDIA3, ENOA, ECH1, PRDX4, ERP29, ECHM), eleven decreased (TRFE, ALBU, K2C1, ACTG, CSH2, PRDX2, FABP5, HBG1, FABP4, K2C8, K1C9) expression in first-trimester compared to the full-term placentas and two proteins exclusively expressed in first-trimester placentas (MESD, MYDGF). Conclusion According to Reactome and PANTHER softwares, these proteins were mostly involved in response to chemical stimulus and stress, regulation of biological quality, programmed cell death, hemostatic and catabolic processes, protein folding, cellular oxidant detoxification, coagulation and retina homeostasis. Elucidation of alteration in protein signature during placental development would provide researchers with a better understanding of the critical biological processes of placentogenesis and delineate proteins involved in regulation of placental function during development.

309. DEVELOPMENT AND VALIDATION OF AN ELISA FOR HIGH TEMPERATURE REQUIREMENT FACTOR A3 (HtrA3): EARLY DETECTION OF PREECLAMPSIA

2010 ◽  
Vol 22 (9) ◽  
pp. 109
Author(s):  
K. Dynon ◽  
G. Nie
Keyword(s):  
Monoclonal Antibodies ◽  
Pregnant Women ◽  
Early Pregnancy ◽  
First Trimester ◽  
High Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Hypertensive Disorder ◽  
Placental Development ◽  
Mortality And Morbidity ◽  
Human Sera

Preeclampsia (PE) is a multisystemic condition in pregnant women that can be life threatening for both mother and baby. PE is a hypertensive disorder that develops concurrently with proteinuria after 20 weeks of gestation. Abnormal placental development during early pregnancy precedes the onset of PE later in gestation. Early diagnosis of PE is essential to reduce PE-related mortality and morbidity. To date there is no clinically useful biochemical diagnostic method that can detect PE during early pregnancy. Our laboratory discovered and cloned the serine protease HtrA3 and has shown that HtrA3 protein levels are intimately involved in placentation (1). Persistently high serum levels of HtrA3 are detected at the end of the first trimester in pregnant women who subsequently develop PE, suggesting that monitoring HtrA3 in maternal blood during early pregnancy may identify women at risk for PE. To develop monoclonal antibodies specific for HtrA3; and to develop an enzyme-linked immunosorbent assay (ELISA) to detect HtrA3 in human sera. Monoclonal antibodies were generated against full length human HtrA3 and small HtrA3 peptides and tested on recombinant HtrA3, human sera and first trimester decidual and villous tissues using western blot, immunoprecipitation and Amplified luminescent proximity homogeneous assay (Alpha)LISA technology. Three antibody pairs were identified that detected either short and/or long isoforms of HtrA3 in sera and placental tissues. Recombinant HtrA3 was detected by AlphaLISA and higher levels of HtrA3 were detected in serum of PE women compared to gestation-matched controls in preliminary testing. These antibody pairs can now be used for the development of specific and high throughput assays. The AlphaLISA will then be used to validate that abnormal serum HtrA3 levels during early pregnancy can predict preeclampsia. (1) Nie et al, (2006) Biol Reprod 74, 366–374.

scholarly journals Ribosomal protein S7 ubiquitination during ER stress in yeast is associated with selective mRNA translation and stress outcome

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Yasuko Matsuki ◽  
Yoshitaka Matsuo ◽  
Yu Nakano ◽  
Shintaro Iwasaki ◽  
Hideyuki Yoko ◽  
...  
Keyword(s):  
Er Stress ◽  
Ribosomal Protein ◽  
Translational Control ◽  
Ribosomal Proteins ◽  
Translational Regulation ◽  
Mrna Translation ◽  
Ribosome Profiling ◽  
Independent Manner ◽  
Eif2α Phosphorylation ◽  
The Unfolded Protein Response

AbstracteIF2α phosphorylation-mediated translational regulation is crucial for global translation repression by various stresses, including the unfolded protein response (UPR). However, translational control during UPR has not been demonstrated in yeast. This study investigated ribosome ubiquitination-mediated translational controls during UPR. Tunicamycin-induced ER stress enhanced the levels of ubiquitination of the ribosomal proteins uS10, uS3 and eS7. Not4-mediated monoubiquitination of eS7A was required for resistance to tunicamycin, whereas E3 ligase Hel2-mediated ubiquitination of uS10 was not. Ribosome profiling showed that the monoubiquitination of eS7A was crucial for translational regulation, including the upregulation of the spliced form of HAC1 (HAC1i) mRNA and the downregulation of Histidine triad NucleoTide-binding 1 (HNT1) mRNA. Downregulation of the deubiquitinating enzyme complex Upb3-Bre5 increased the levels of ubiquitinated eS7A during UPR in an Ire1-independent manner. These findings suggest that the monoubiquitination of ribosomal protein eS7A plays a crucial role in translational controls during the ER stress response in yeast.

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