Human mesothelial cells (LP9/TERT-1) were exposed to low and high (15 and 75 m2/cm2 dish) equal surface area concentrations of crocidolite asbestos, nonfibrous talc, fine titanium dioxide (TiO2), or glass beads for 8 or 24 h. RNA was then isolated for Affymetrix microarrays, GeneSifter analysis and QRT-PCR. Gene changes by asbestos were concentration- and time-dependent. At low nontoxic concentrations, asbestos caused significant changes in mRNA expression of 29 genes at 8 h and 205 genes at 24 h, whereas changes in mRNA levels of 236 genes occurred in cells exposed to high concentrations of asbestos for 8 h. Human primary pleural mesothelial cells also showed the same patterns of increased gene expression by asbestos. Nonfibrous talc at low concentrations in LP9/TERT-1 mesothelial cells caused increased expression of 1 gene Activating Transcription Factor 3 (ATF3) at 8 h and no changes at 24 h, whereas expression levels of 30 genes were elevated at 8 h at high talc concentrations. Fine TiO2 or glass beads caused no changes in gene expression. In human ovarian epithelial (IOSE) cells, asbestos at high concentrations elevated expression of 2 genes (NR4A2, MIP2) at 8 h and 16 genes at 24 h that were distinct from those elevated in mesothelial cells. Since ATF3 was the most highly expressed gene by asbestos, its functional importance in cytokine production by LP9/TERT-1 cells was assessed using siRNA approaches. Results reveal that ATF3 modulates production of inflammatory cytokines (IL-1, IL-13, G-CSF) and growth factors (VEGF and PDGF-BB) in human mesothelial cells.
Alterations in gene expression in human mesothelial cells correlate with mineral pathogenicity.
Specimen part, Cell line
View SamplesWe have derived induced porcine pluripotent stem cells (iPPSCs) from porcine fetal fibroblasts by lentiviral transduction of four human (h) reprogramming genes, hOCT4, hSOX2, hKLF4 and hc-MYC , the same combination of factors used for deriving induced pluripotent stem cell (iPSC) lines in both mouse and human. The obtained iPPSC lines resemble human embryonic stem cells (ESC) in their gross morphology and dependence on FGF2, on the other hand, the iPPSCs share characteristics like growth rate and cell surface markers with mESC . Additionally, the iPPSCs express pluripotency- associated genes similar to mouse and human iPSCs as well as ESC, along with the pig epiblast cells. Some of the iPPSC lines retained a stable karyotype and phenotype even in culture for a prolonged period of time (passage 39). The iPPSCs can be induced to differentiate along lineages representative of the three embryonic germ layers both in vitro and in vivo demonstrating the pluripotency of these cells.
Derivation of induced pluripotent stem cells from pig somatic cells.
Specimen part
View SamplesThe pig is important for agriculture and as an animal model in human and veterinary medicine, yet, despite over 20 years of effort, it has proved a difficult species from which to generate pluripotent stem cells analogous to those derived from mouse embryos. Here we report the production of LIF-dependent, so called nave type, pluripotent stem cells from the inner cell mass of porcine blastocysts by up-regulating expression of KLF4 and POU5F1. These cells resemble mouse ES cells and are distinct from the FGF2-dependent, induced pluripotent cell type derived from porcine somatic cells.
Leukemia inhibitory factor (LIF)-dependent, pluripotent stem cells established from inner cell mass of porcine embryos.
Sex
View SamplesPurpose: 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. Overall design: 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.
Comparison of syncytiotrophoblast generated from human embryonic stem cells and from term placentas.
No sample metadata fields
View SamplesWe aimed to investigate the function of syndecan-1 in tumor cell adhesion and migration, with special focus on the importance of its distinct protein domains, to better understand the structure-function relationship of syndecan-1 in tumor progression. We utilized two mesenchymal tumor cell lines which were transfected to stably overexpress full-length syndecan-1 or truncated variants: the 78 which lacks the extracellular domain except the DRKE sequence proposed to be essential for oligomerization, the 77 which lacks the whole extracellular domain, and the RMKKK which serves as a nuclear localization signal. Various bioassays for cell adhesion, chemotaxis, random movement and wound healing were studied. Furthermore we performed gene microarray to analyze the global gene expression pattern influenced by syndecan-1.
Novel genes and pathways modulated by syndecan-1: implications for the proliferation and cell-cycle regulation of malignant mesothelioma cells.
No sample metadata fields
View SamplesThe transcriptomic responses of syndecan-1 silencing in a human mesothelioma cell line was followed with microarray analysis. To project the transcriptome analysis on the full-dimensional picture of cellular regulation, we applied a novel method of network enrichment analysis which elucidated signalling relations between differentially expressed genes and pathways acting via various molecular mechanisms.
Novel genes and pathways modulated by syndecan-1: implications for the proliferation and cell-cycle regulation of malignant mesothelioma cells.
Cell line
View SamplesHuman pluripotent stem cells (hPSC) exposed to BMP4 (B) and inhibitors of ACTIVIN signaling (A83-01; A) and FGF2 (PD173074; P) in absence of FGF2 (BAP conditions) differentiate into colonies primarily comprised of trophoblast. In an attempt to isolate trophoblast stem cells, colonies of hESC were exposed to BAP for 24 h at which time they had begun to transition into a CDX2-positive state. Cultures were then dissociated into single cells by trypsin and grown on a gelatin substratum. Under these conditions, organized CDX2+/KRT7- colonies began to emerge within a few days. The self-renewing cell lines were not TBSC, but met standard criteria for pluripotency. They were named H1BP cells. They differed from the progenitor hPSC in morphology, ability to be clonally propagated from single cells onto gelatin, requirements for FGF2, and transcriptome profile.
Heightened potency of human pluripotent stem cell lines created by transient BMP4 exposure.
Specimen part, Time
View SamplesThis SuperSeries is composed of the SubSeries listed below.
Specificity and heterogeneity of terahertz radiation effect on gene expression in mouse mesenchymal stem cells.
Specimen part
View SamplesWe report that terahertz (THz) irradiation of mouse mesenchymal stem cells with a pulsed broadband (centered at 10 THz) source, or a single-frequency, 2.52 THz, (SF) laser source, both with weak average power (<1mW/cm2), results in specific heterogenic changes in gene expression. The insignificant differential expression of heat shock and stress related genes as well as our temperature measurements imply a non-thermal response. The microarray survey and RT-PCR experiments demonstrate that at different irradiation conditions distinct groups of genes are activated. Stem cells irradiated for 12 hours with the broadband THz source exhibit an accelerated differentiation toward adipose phenotype, while the 2-hour (broadband or SF) irradiation affects genes transcriptionally active in pluripotent stem cells. Phenotypic and gene expression differences suggest that the THz effect depends on irradiation parameters such as duration and type of THz source, and on the level of stem cell differentiation. Computer simulations of the core promoters of two pluripotency markers reveal association between gene upregulation and propensity for DNA breathing. We propose that THz radiation has potential for non-contact control of cellular gene expression.
Specificity and heterogeneity of terahertz radiation effect on gene expression in mouse mesenchymal stem cells.
Specimen part
View SamplesWe report that terahertz (THz) irradiation of mouse mesenchymal stem cells with a pulsed broadband (centered at 10 THz) source, or a single-frequency, 2.52 THz, (SF) laser source, both with weak average power (<1mW/cm2), results in specific heterogenic changes in gene expression. The insignificant differential expression of heat shock and stress related genes as well as our temperature measurements imply a non-thermal response. The microarray survey and RT-PCR experiments demonstrate that at different irradiation conditions distinct groups of genes are activated. Stem cells irradiated for 12 hours with the broadband THz source exhibit an accelerated differentiation toward adipose phenotype, while the 2-hour (broadband or SF) irradiation affects genes transcriptionally active in pluripotent stem cells. Phenotypic and gene expression differences suggest that the THz effect depends on irradiation parameters such as duration and type of THz source, and on the level of stem cell differentiation. Computer simulations of the core promoters of two pluripotency markers reveal association between gene upregulation and propensity for DNA breathing. We propose that THz radiation has potential for non-contact control of cellular gene expression.
Specificity and heterogeneity of terahertz radiation effect on gene expression in mouse mesenchymal stem cells.
Specimen part
View Samples