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Next Generation Sequencing of syncytializing first trimester human cytotrophoblasts.

ABSTRACT: Aim of this study was to monitor differentially expressed gene signatures of human first trimester villous cytotrophoblasts (vCTBs) undergoing spontaneous differentiation into syncytiotrophoblasts (STB).

ORGANISM(S): Homo sapiens

PROVIDER: GSE143860 | GEO | 2020/05/19

REPOSITORIES: GEO

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Project description:Invasion of cytotrophoblasts into uterine tissues is essential for placental development. To identify molecules regulating trophoblast invasion, mRNA signatures of purified villous (CTB, poor invasiveness) and extravillous (EVT, high invasiveness) trophoblasts isolated from first trimester human placentae and villous explant cultures, respectively, were compared using GeneChip analyses yielding 991 invasion/migration related transcripts. Several genes involved in physiological and pathologic cell invasion, including ADAM-12,-19,-28 as well as Spondin-2, were upregulated in EVT. Pathway prediction analyses identified several functional modules associated with either the invasive or the non-invasive trophoblast phenotype. One of the genes which were downregulated in the invasive mRNA pool, heme oxygenase-1 (HO-1), was selected for functional analyses. Real-time PCR analyses, Western blottting, and immunofluorescene of first trimester placentae and differentiating villous explant cultures demonstrated downregulation of HO-1 in invasive EVT as compared to CTB. Modulation of HO-1 expression in loss-of as well as gain-of function cell models (BeWo and HTR8/SVneo, respectively) demonstrated an inverse relationship of HO-1 expression with trophoblast migration in transwell and wound healing assays. Importantly, HO-1 expression led to an increase in protein levels and activity of the nuclear hormone receptor PPARgamma. Pharmacological inhibition of PPARgamma abrogated the inhibitory effects of HO-1 on trophoblast migration. Collectively, our results demonstrate that gene expression profiling of EVT and CTB can be used to unravel novel regulators of cell invasion. Accordingly, we identify heme oxygenase-1 as a negative regulator of trophoblast motility acting via upregulation of PPARgamma. Experiment Overall Design: To identify genes potentially regulating cell invasion trophoblast cells of early human gestation with distinct invasive properties were profiled. Experiment Overall Design: Distinct gene expression signatures of highly invasive EVT (n = 6) and poorly invasive CTB (n = 5) of different first trimester placentae using Affymetrix U133A GeneChips interrogating >20,000 genes were determined.

Project description:Oxidative stress plays a crucial role in preeclampsia (PE) pathogenesis. Evidence indicates altered microRNAs (miRNAs) expression in PE, however, the relationship between oxidative stress and miRNA changes in placenta as a potential mechanism involved in PE is not fully elucidated. In the present study, we investigate the impact of increased oxidative stress on miRNA and mRNA expression profiles of genes known to be associated with PE in villous 3A first trimester trophoblast cells. Cells were exposed to H2O2 at 10 different concentrations (0-1 mM) for 0.5, 4, 24, and 48 h. Cytotoxicity was determined using the SRB assay and data was used to calculate the IC50 of H2O2. Total RNA was extracted after short-term exposure (4 h) to H2O2 for miRNA expression profiling. H2O2 exerted a concentration and time-dependent cytotoxicity on 3A trophoblast cells. Short-term exposure of 3A cells to low concentration of H2O2 (5% of IC50) significantly altered miRNA profile as evidenced by significant changes in 195 out of 595 evaluatable miRNAs. Short-term exposure of villous first trimester trophoblasts to low concentrations of H2O2 significantly alters miRNA profile. Oxidative stress plays a crucial role in preeclampsia (PE) pathogenesis. Evidence indicates altered microRNAs (miRNAs) expression in PE, however, the relationship between oxidative stress and miRNA changes in placenta as a potential mechanism involved in PE is not fully elucidated. In the present study, we investigate the impact of increased oxidative stress on miRNA and mRNA expression profiles of genes known to be associated with PE in villous 3A first trimester trophoblast cells. Cells were exposed to H2O2 at 10 different concentrations (0-1 mM) for 0.5, 4, 24, and 48 h. Cytotoxicity was determined using the SRB assay and data was used to calculate the IC50 of H2O2. Total RNA was extracted after short-term exposure (4 h) to H2O2 for miRNA expression profiling. H2O2 exerted a concentration and time-dependent cytotoxicity on 3A trophoblast cells. Short-term exposure of 3A cells to low concentration of H2O2 (5% of IC50) significantly altered miRNA profile as evidenced by significant changes in 195 out of 595 evaluatable miRNAs. Short-term exposure of villous first trimester trophoblasts to low concentrations of H2O2 significantly alters miRNA profile.

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Next Generation Sequencing of syncytializing first trimester human cytotrophoblasts.

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