Comparison of syncytiotrophoblast differentiated from human pluripotent stem cells and that derived from primary cytotrophoblast recovered from term placentae
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Toshihiko Ezashi
PROVIDER: E-GEOD-73017 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:We compare two groups of transcriptome profiles. They are 1) primary human trophoblasts (PHT) and 2) syncytiotrophoblast from human pluripotent stem cells. This data set represents the former data set and the latter has been deposited separately at GSE72712. Primary human trophoblasts (PHT) were purified cytotrophoblast populations from whole term placentas and then cultured in vitro. The cytotrophoblasts were cultured for short (9 h) or long (48 h) term that show undifferentiated (PHTu) or differentiated (PHTd) phenotype that led syncytialization, respectively. Transcriptome profiles of PHTu and PHTd were compared with three size fractioned syncytiotrophoblasts and trophoblasts from human pluripotent stem cells that are named ESCd_gt70 (greater than 70 µm), ESCd_40-70 (size between 40 and 70 µm), ESCd_lt40 (smaller than 40 µm) and undifferentiated progenitor (ESCu; see GSE72712). Primary human trophoblasts (PHT) were derived and cultured from three term human placentas obtained by the Obstetrical Specimen Procurement Unit from women after a healthy pregnancy, labor, and delivery at Magee-Womens Hospital of the University of Pittsburgh Medical Center. Isolated PHT lines obtained from three individual (1 female and 2 male) placentas were cultured separately.
2016-04-06 | E-GEOD-73016 | biostudies-arrayexpress
Project description:Purpose: Syncytiotrophoblast (STB) is a multi-nucleated, terminally differentiated syncytium that covers the surface of the villous placenta and forms the major interface with maternal blood. It releases placental hormones and plays a primary role in exchange of gases, nutrients and waste products. Alterations in STB development and turnover have been implicated in placental diseases, including preeclampsia (PE). In vitro cell models are badly needed to study STB development and physiology due to inaccessibility to placental tissues during gestation. To establish in vitro STB model system, we generate STB and its mononucleated precursors from human embryonic stem cells (hESC) and profile for RNA content by RNAseq. Methods: H1 Human ESC (WA01) were treated with BMP4, the ALK4/5/7 inhibitor (A83-01), and the FGF2 signaling inhibitor (PD173074) (BAP) to direct them to the trophoblast lineage and provided both STB and extravillous trophoblast. Syncytial areas emerged at day 8 BAP treatment ranged in diameter from ~40 µm to > 100 µm. The intact syncytial areas were isolated by sieving successively through 70 μm and 40 μm mesh cell strainers. The captured cells are recovered by inverting the strainer and rinsing with culture medium to separate large (>70 μm) and middle size cell sheets (40-70 μm). The fraction that passes through both sieves represents cells of smallest diameter (< 40 μm), presumably cytotrophoblast. Total 12 RNA samples from triplicate three size-fractioned BAP treated and three untreated hESC cultured in a FGF2 supplemented medium in parallel were analyzed. Results: The larger > 70 μm areas stained positively for STB markers while ultrastructural analysis clearly revealed multi-nuclear cells with an extensive cytoplasm containing many prominent secretion granules. The larger STB areas also had a larger DNA content that > 70 μm fraction contained 37 times more nuclear content and 40-70 µm fraction did 16 times more. Compared to the < 40 μm cell fraction, these larger cells over-expressed a full repertoire of genes characteristic of STB, e.g. CGA, CGB, PGF, ERVW1, GCM1. The smallest cell fraction had a DNA content consistent with mononuclear diploid cells, contained few secretory granules, and were only weakly positive for STB markers. Conclusion: The data are consistent with the > 70 μm cells being mature STB, while the intermediate fraction may represent a precursor population. Human ESC directed along the trophoblast lineage by BAP treatment offers a useful model for following STB formation in vitro and suggest that this protocol may have utility in studying the basis of certain placental diseases, especially preeclampsia, where placental tissue isolated at term or after pregnancy terminations can only offer limited information. Three size fraction mRNA profiles of syncytial areas emerged at day 8 BAP treatment of hESC were generated by deep sequencing along with untreated hESC, in triplicate, using Illumina HiSeq 2500.
2016-04-06 | E-GEOD-72712 | biostudies-arrayexpress
Project description:Low back pain is a major cause of disability especially for people between 20 and 50 years of age. As a costly healthcare problem, it imposes a serious socio-economic burden. Current surgical therapies have considerable drawbacks and fail to replace the normal disc in facilitating spinal movements and absorbing load. Therefore, the focus of regenerative medicine is on identifying biomarkers and signalling pathways to improve our understanding about the cascades of disc degeneration and allow for the design of specific therapies. We hypothesized that comparing microarray profiles from degenerative and non-degenerative discs will lead to the identification of dysregulated signalling and pathophysiological targets. Microarray data sets were generated from human annulus fibrosus cells and analysed using IPA ingenuity pathway analysis system. Gene expression values were validated by qRT-PCR, and respective proteins were identified by immunohistochemistry. Microarray analysis revealed 17 dysregulated molecular markers and various dysregulated cellular functions, including cell proliferation and inflammatory response, in the human degenerative annulus fibrosus. The most significant canonical pathway induced in degenerative annulus fibrosus was found to be the interferon signalling pathway. In conclusion, this study indicates interferon-alpha signalling pathway activation with IFIT3 and IGFBP3 up-regulation which may affect cellular function in human degenerative disc. 48 samples of intervertebral disc tissue - annulus fibrosus and nucleus pulposus - displaying varying degrees (grades) of degeneration
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Project description:Proteomics assessment of supernatants, cell contents of trophoblasts were performed to identify what proteins are secreted from this particular placental region.
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Project description:YY1 is a sequence-specific DNA-binding transcription factor that has many important biological roles. However, its function in trophoblasts at the maternal-foetal interface remains to be elucidated. In this study, we used an mRNA microarray and quantitative reverse transcription-PCR and compared the YY1 mRNA expression level in trophoblasts between patients with recurrent miscarriage (RM) and healthy control subjects. Our results revealed that YY1 mRNA expression was significantly lower in the trophoblasts of the RM group compared with the healthy control group. Furthermore, immunofluorescence and immunohistochemical data showed that YY1 was highly expressed in human placental villi during early pregnancy, especially in cytotrophoblast cells and invasive extravillous trophoblasts, and it was expressed at a much lower level in the placental villi of term pregnancy. YY1 overexpression enhanced the invasion and proliferation of trophoblasts, while knockdown of YY1 repressed these effects. Antibody array screening revealed that YY1 significantly promoted MMP2 expression in trophoblasts. Bioinformatics analysis identified three YY1-binding sites in the MMP2 promoter region, and chromatin immunoprecipitation analysis verified that YY1 binds directly to its promoter region. Importantly, inhibition of YY1 by siRNA clearly decreased trophoblast invasion in an ex vivo explant culture model. Overall, our findings revealed a new regulatory pathway of YY1/MMP2 in trophoblast cells invasion during early pregnancy, and indicated that YY1 may be involved in the pathogenesis of RM. Total RNA was isolated using Trizol from trophoblast cells from three healthy controls (HC) and three recurrent miscarriage (RM) patients. Total RNA were extracted and used for hybridizing Affymetrix chips (GeneChip® Human Transcriptome Array 2.0(HTA2.0)). Data were normalised by gcRMA method and raw p-values adjusted by Bonferroni procedure (1%).
2016-01-21 | E-GEOD-76862 | biostudies-arrayexpress
Project description:Analyze gene expression levels in primary trophoblasts, derived from term human placenta and cultured under standard or hypoxic conditions Human placental trophoblasts were dispersed using a trypsin-deoxyribonuclease-dispase/Percoll method, plated in 6-well plates, and maintained in standard culture conditions (O2=20%). After 4 h (defined as time 0), the plates were divided to those in continued standard culture conditions, or to culture in hypoxia (O2=0%). Cells were then harvested at 6 h, 12 h, 24 h, 48 h and 72 h, and processed for mRNA arrays
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Project description:Analyze gene expression profiles in HTR-8/SVeo cells stably transfected with a BAC plasmid containing the entire C19MC miRNA cluster HTR-8/SVeo cells were transfected with a modified BAC plasmid containing the entire C19MC miRNA cluster and carrying a zeocin selection cassette. Several independent clones and a mixed population of transfected cells were analyzed and compared to non-transfected HTR-8/SVeo cells and primary human trophoblasts that express the C19MC miRNAs endogenously
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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
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