Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: PEG10 mRNA expression in placental lysates from patients with early-onset preeclampsia. PEG10 mRNA expression was reduced in placentas from 79 women with early-onset preeclampsia (pink line) compared to 18 gestation-matched preterm controls (grey) ( A ). Further analysis of the early-onset preeclampsia cohort (blue circle) and gestation-matched pre-term controls (grey diamond) showed no significant association between gestation ( B ) or birth weight ( C ). Data expressed as a median with interquartile range. Simple linear regression was used to present a line of best fit (solid line) and 95% confidence interval (dashed line). Individual symbols represent individual patients. P * < 0.05

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Expressing

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: Localisation of PEG10 protein and mRNA within the placenta. Immunohistochemistry (IHC) staining of human placental villi shows PEG10 protein (brown) localised to cytotrophoblasts in both control and early-onset preeclampsia tissue ( A ). Semi-quantitative, blinded assessment of IHC staining indicated reduced PEG10 protein in early-onset preeclampsia placentas relative to controls ( C ). In situ hybridisation shows PEG10 (blue) and PECAM-1 (pink) mRNA expression. PEG10 expression localised to cytotrophoblasts in both control and early-onset preeclampsia tissues while PECAM-1 marked placental vasculature ( B ). No staining observed in respective isotype controls (minimised image). Figures were captured at 40 × magnification and are representative of 5 experimental repeats. Data expressed as mean \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\pm$$\end{document} ± SEM. Individual symbols represent individual scores and P * < 0.05

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Immunohistochemistry, Control, In Situ, Hybridization, Expressing, Staining

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: PEG10 gene expression reduces with cytotrophoblast differentiation. First-trimester human cytotrophoblast stem cells (hTSCs) were differentiated into syncytiotrophoblasts (ST) over 96 h ( A-D ) . Effective syncytialisation was confirmed by a decrease in trophoblast markers, TEAD4 (A) and CDH2 ( B ) and an increase in ST marker, SDC1 ( C ). PEG10 mRNA expression significantly decreased by 96 h in ST cells ( D ). hTSCs were differentiated into extravillous trophoblast (EVT) cells across 144 h as shown by an increase in EVT marker, HLAG ( E ). PEG10 mRNA expression was significantly decreased after 144 h of EVT differentiation ( F ). Data expressed as a median with interquartile range. Individual symbols represent n = 5 experimental repeats. P ** < 0.01, P *** < 0.001

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Gene Expression, Marker, Expressing

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: The effect of hypoxia and inflammatory cytokines on PEG10 expression in cytotrophoblast stem cells. First-trimester human cytotrophoblast stem cells (hTSCs) or term primary cytotrophoblast were exposed to ( A-B ). PEG10 mRNA expression was significantly reduced in hTSCs exposed to hypoxia (1% oxygen) relative to normoxia (8% oxygen) ( A ). A similar finding was observed when primary term trophoblast were exposed to hypoxia ( B ). Next, hTSCs were treated with increasing doses of TNFα (0, 0.1, 1 and 10 ng/mL) and showed a dose-dependent reduction in PEG10 expression when compared to vehicle-treated controls ( C ). No changes in PEG10 expression were observed in cells with increasing doses (0, 0.1, 1, 10 ng/mL) of IL-6 ( D ). Data is presented as a median with interquartile range. Individual symbols represent n = 5 experimental repeats and P ** < 0.01

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Expressing

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: Effect of silenced PEG10 on trophoblast differentiation. First-trimester human cytotrophoblast stem cells (hTSCs) were treated with PEG10 siRNA prior to differentiation into syncytiotrophoblasts over 96 h. No change in the expression of hTSC and syncytiotrophoblast markers, CDH2 ( A ) and SDC1 ( B ) were observed in PEG10 siRNA-treated cells, relative to negative controls. Next, PEG10-silenced hTSCs were differentiated into EVTs over 144 h. No change in hTSC or EVT markers, TEAD4 ( C ) and HLAG ( D ) were observed in PEG10 siRNA-treated cells, compared to controls. Data expressed as a median with interquartile range. Individual symbols represent n = 5 experimental repeats

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Expressing

Journal: Reproductive Biology and Endocrinology : RB&E

Article Title: Paternal Expressed Gene 10 (PEG10) is decreased in early-onset preeclampsia

doi: 10.1186/s12958-023-01116-3

Figure Lengend Snippet: Effect of silenced PEG10 on cytotrophoblast function. Following transfection with luciferase reporter plasmid containing four copies of the SMAD binding element, PEG10 -siRNA hTSCs had reduced basal levels of canonical TGF \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$-\beta$$\end{document} - β signalling relative to negative siRNA controls ( A ). Real-time analysis of cell proliferation showed no effect of PEG10 siRNA (pink) relative to control siRNA treated cells (black) ( B ). Dotted line in panel B indicates time of siRNA treatment, dashed line indicates time of media change. Data expressed as median with interquartile range. P * < 0.05

Article Snippet: Taqman Fast Advanced Master Mix (ThermoFisher Scientific, Waltham, USA) and specific fluorescein amidite (FAM)-labelled Taqman gene expression assays (Life Technologies, Carlsbad, USA) were used to measure mRNA expression of PEG10 (Assay ID: Hs01122880_m1), CDH2 (n-cadherin 2, Assay ID: Hs00983056_m1), GATA3 (GATA binding protein 3, Assay ID: Hs00231122_m1), HLAG (human leukocyte antigen G, Assay ID: Hs03045108_m1), SDC1 (syndecan-1, Assay ID: Hs00896423_m1) and TEAD4 (TEA domain transcription factor 4, Assay ID: Hs01125032_m1).

Techniques: Transfection, Luciferase, Plasmid Preparation, Binding Assay, Control

Journal: Molecular reproduction and development

Article Title: Novel key roles for Structural maintenance of chromosome flexible domain containing 1 (Smchd1) during preimplantation mouse development

doi: 10.1002/mrd.23001

Figure Lengend Snippet: Primers for qPCR

Article Snippet: Rpl18 was selected for later studies at the morula stage examining effects of siRNA knockdown on the expression of mRNAs identified as affected in the RNAseq analysis, as RNAseq data revealed it to be highly constant in expression level with minimal variance at that stage. table ft1 table-wrap mode="anchored" t5 Table 2. caption a7 Target Assay Sequence/Catalog # Primer Efficiency (%) Arid3a Taqman Mm00492248_m1 96.6 β-Actin Syber Green F-CTGAACCCTAAGGCCAACC R-CCAGAGGCATACAGGGACAG Cdx2 Syber Green F-AGACAAATACCGGGTGGTGTA R-CCAGCTCACTTTTCCTCCTGA Dppa1 Taqman Mm00626454_m1 92.9 Eomes Syber Green F-TGTGACGGCCTACCAAAACA R-GCCGTGTACATGGAATCGTAG Igf2r Taqman Mm00439576_m1 96.5 Krt8 Syber Green F-CTTCATTGACAAGGTGCGCTT R-TTGCTCCTCGACGTCTTCTG Nt5c3b Taqman Mm00518281_m1 92.4 Peg10 Taqman Mm01167724_m1 100 Rpl18 Taqman Mm02745785_g1 100 Skp2 Taqman Mm00449925_m1 97.6 Smchd1 Taqman Mm00512253_m1 89.3 Smchd1 Syber Green F-TACTGACGGCGACGAAAGAAA R-CTCTGCAAGAGCAAATGGCAAA Ubtf Taqman Mm00456972_m1 93.9 Open in a separate window Primers for qPCR Other studies used embryos from Cdx2 +/− inter se matings to test for effects of Cdx2 mutation on Smchd1 expression.

Techniques: Sequencing

Journal: Molecular reproduction and development

Article Title: Novel key roles for Structural maintenance of chromosome flexible domain containing 1 (Smchd1) during preimplantation mouse development

doi: 10.1002/mrd.23001

Figure Lengend Snippet: Genes differentially expressed in Smchd1 siRNA knockdown embryos

Article Snippet: Rpl18 was selected for later studies at the morula stage examining effects of siRNA knockdown on the expression of mRNAs identified as affected in the RNAseq analysis, as RNAseq data revealed it to be highly constant in expression level with minimal variance at that stage. table ft1 table-wrap mode="anchored" t5 Table 2. caption a7 Target Assay Sequence/Catalog # Primer Efficiency (%) Arid3a Taqman Mm00492248_m1 96.6 β-Actin Syber Green F-CTGAACCCTAAGGCCAACC R-CCAGAGGCATACAGGGACAG Cdx2 Syber Green F-AGACAAATACCGGGTGGTGTA R-CCAGCTCACTTTTCCTCCTGA Dppa1 Taqman Mm00626454_m1 92.9 Eomes Syber Green F-TGTGACGGCCTACCAAAACA R-GCCGTGTACATGGAATCGTAG Igf2r Taqman Mm00439576_m1 96.5 Krt8 Syber Green F-CTTCATTGACAAGGTGCGCTT R-TTGCTCCTCGACGTCTTCTG Nt5c3b Taqman Mm00518281_m1 92.4 Peg10 Taqman Mm01167724_m1 100 Rpl18 Taqman Mm02745785_g1 100 Skp2 Taqman Mm00449925_m1 97.6 Smchd1 Taqman Mm00512253_m1 89.3 Smchd1 Syber Green F-TACTGACGGCGACGAAAGAAA R-CTCTGCAAGAGCAAATGGCAAA Ubtf Taqman Mm00456972_m1 93.9 Open in a separate window Primers for qPCR Other studies used embryos from Cdx2 +/− inter se matings to test for effects of Cdx2 mutation on Smchd1 expression.

Techniques: Knockdown, Binding Assay, Sequencing

Journal: Molecular reproduction and development

Article Title: Novel key roles for Structural maintenance of chromosome flexible domain containing 1 (Smchd1) during preimplantation mouse development

doi: 10.1002/mrd.23001

Figure Lengend Snippet: Quantitative RT-PCR analysis of mRNA expression changes with Smchd1 siRNA knockdown at morula and blastocyst stages.

Article Snippet: Rpl18 was selected for later studies at the morula stage examining effects of siRNA knockdown on the expression of mRNAs identified as affected in the RNAseq analysis, as RNAseq data revealed it to be highly constant in expression level with minimal variance at that stage. table ft1 table-wrap mode="anchored" t5 Table 2. caption a7 Target Assay Sequence/Catalog # Primer Efficiency (%) Arid3a Taqman Mm00492248_m1 96.6 β-Actin Syber Green F-CTGAACCCTAAGGCCAACC R-CCAGAGGCATACAGGGACAG Cdx2 Syber Green F-AGACAAATACCGGGTGGTGTA R-CCAGCTCACTTTTCCTCCTGA Dppa1 Taqman Mm00626454_m1 92.9 Eomes Syber Green F-TGTGACGGCCTACCAAAACA R-GCCGTGTACATGGAATCGTAG Igf2r Taqman Mm00439576_m1 96.5 Krt8 Syber Green F-CTTCATTGACAAGGTGCGCTT R-TTGCTCCTCGACGTCTTCTG Nt5c3b Taqman Mm00518281_m1 92.4 Peg10 Taqman Mm01167724_m1 100 Rpl18 Taqman Mm02745785_g1 100 Skp2 Taqman Mm00449925_m1 97.6 Smchd1 Taqman Mm00512253_m1 89.3 Smchd1 Syber Green F-TACTGACGGCGACGAAAGAAA R-CTCTGCAAGAGCAAATGGCAAA Ubtf Taqman Mm00456972_m1 93.9 Open in a separate window Primers for qPCR Other studies used embryos from Cdx2 +/− inter se matings to test for effects of Cdx2 mutation on Smchd1 expression.

Techniques: Quantitative RT-PCR, Expressing, Knockdown