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Genome-wide expression profile of first trimester villous and extravillous human trophoblast cells.

ABSTRACT: We have examined the transcriptional changes associated with differentiation from villous to extravillous trophoblast using a whole genome microarray. Villous trophoblast (VT) is in contact with maternal blood and mediates nutrient exchange whereas extravillous trophoblast (EVT) invades the decidua and remodels uterine arteries. Using highly purified first trimester trophoblast we identified over 3000 transcripts that are differentially expressed. Many of these transcripts represent novel functions and pathways that show co-ordinated up-regulation in VT or EVT. In addition we identify new players in established functions such as migration, immune modulation and cytokine or angiogenic factor secretion by EVT. The transition from VT to EVT is also characterised by alterations in transcription factors such as STAT4 and IRF9, which may co-ordinate these changes. Transcripts encoding several members of the immunoglobulin-superfamily, which are normally expressed on leukocytes, were highly transcribed in EVT but not expressed as protein, indicating specific control of translation in EVT. Interactions of trophoblast with decidual leukocytes are involved in regulating EVT invasion. We show that decidual T-cells, macrophages and NK cells express the inhibitory collagen receptor LAIR-1 and that EVT secrete LAIR-2, which can block this interaction. This represents a new mechanism by which EVT can modulate leukocyte function in the decidua. Since LAIR-2 is detectable in the urine of pregnant, but not non-pregnant women, trophoblast-derived LAIR-2 may also have systemic effects during pregnancy.

SUBMITTER: Apps R

PROVIDER: S-EPMC3065343 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Genome-wide expression profile of first trimester villous and extravillous human trophoblast cells.

Apps R R Sharkey A A Gardner L L Male V V Trotter M M Miller N N North R R Founds S S Moffett A A

Placenta 20101113 1

We have examined the transcriptional changes associated with differentiation from villous to extravillous trophoblast using a whole genome microarray. Villous trophoblast (VT) is in contact with maternal blood and mediates nutrient exchange whereas extravillous trophoblast (EVT) invades the decidua and remodels uterine arteries. Using highly purified first trimester trophoblast we identified over 3000 transcripts that are differentially expressed. Many of these transcripts represent novel functi ...[more]

PMID: 21075446

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Project description:Purpose: During early placentation, extravillous trophoblast (EVT) cell invasion into maternal decidua plays a crucial role in establishment of the successful pregnancy. However, little is known about the characteristic features of EVT-associated ncRNAs. The purpose of this study is to elucidate gene expression profile for both coding and non-coding genes (i.e., mRNAs, lncRNAs, and miRNAs) expressed in EVT cells isolated from the first-trimester human placenta by RNA sequencing analysis. Methods: Human first trimester placental tissues (at gestational week 7, n=3) were obtained after legal abortions. EVT cells growing from explanted human chorionic villi were isolated. Minced chorionic villous tips were defined as first-trimester chorionic villous trophoblast (CVT) cells. Using total RNAs of these samples, mRNA/lncRNA-sequencing analysis was carried out on an Illumina MiSeq (San Diego, California, USA) using MiSeq v2 Reagent Kit 300cycle (cat. no. MS-102-2002; Illumina) and MiSeq v2 Reagent Kit 50cycle (cat. no. MS-102-2001; Illumina), respectively. The paired-end reads of 151 bp length for mRNA/lncRNA-sequencing and single reads of 26 bp length for miRNA-sequencing were obtained as fastq files which were imported into CLC genomics workbench (version 8.0.1; Qiagen, Venlo, Netherlands). Clean reads were aligned to the human genome sequence retrieved from the Ensembl database (Assembly GRCh38.p13, database version 82.38 and 101.38) and microRNA database miRbase (release 21), using CLC Genomics Workbench Map Reads to Reference tool. Normalization of gene expression data was performed by the TMM method of edgeR package. Statistically differentially expressed transcripts were conducted using edgeR. Genes that had a false discovery rate (FDR)-corrected p-value (q-value) of < 0.05 and an absolute log2 fold change (log FC) ≥ 1 were judged to be significantly differentially expressed (i.e., differentially expressed genes, DEGs). Results: Five thousand ninty-two mRNAs , 422 lncRNAs and 400 miRNAs were found to be differentially expressed. One hundred differentially expressed genes (i.e., 10 mRNAs, 7 lncRNAs, and 83 miRNAs) that are located in the chromosome 14q32.2 region containing the eutherian-specific imprinted DLK1-DIO3 domain were significantly down-regulated in EVT cells. In terms of imprinted genes (http://www.geneimprint.com/site/genes-by-species), 68 genes were found to be differentially expressed; 71% of these genes were down-regulated in EVT cells. lncRNA H19 was a DEG (q = 1.29.E-09, log FC = 2.98) and the most highly expressed lncRNA (70.47%) in EVT cells. Conclusions: The present research should be valuable for a resource for future ncRNA research in EVT cells.

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.

Dataset Information

Genome-wide expression profile of first trimester villous and extravillous human trophoblast cells.

Publications

Genome-wide expression profile of first trimester villous and extravillous human trophoblast cells.

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