The use of human pluripotent stem cells (hPSCs) in cell therapy is hindered by the tumorigenic risk from residual undifferentiated cells. Here we performed a high-throughput screen of over 52,000 small molecules, and identified 15 highly selective cytotoxic inhibitors of hPSCs (PluriSIns). Cellular and molecular analyses revealed that the most selective compound, PluriSIn #1, is a pluripotent-specific inhibitor of stearoyl-coA desaturase (SCD1), the key enzyme in the biosynthesis of monounsaturated fatty acids (MUFA). SCD1 inhibition in hPSCs induced ER stress, protein synthesis attenuation, and apoptosis of these cells, revealing that MUFA biosynthesis is crucial for their survival. PluriSIn #1 was also cytotoxic toward the ICM cells of mouse embryos, indicating that the dependence on SCD1 is inherent to the pluripotent state. Finally, application of PluriSIn #1 prevented teratoma formation from tumorigenic undifferentiated cells. Our novel method to eliminate undifferentiated cells from culture should thus increase the safety of hPSC-based treatments.
Selective elimination of human pluripotent stem cells by an oleate synthesis inhibitor discovered in a high-throughput screen.
Specimen part, Cell line, TreatmentView Samples
We report how the zebrafish melanoma cell line ZMEL1 changes after intravascular injection into 2dpf zebrafish embryos, as compared to the cells growing in vitro. Overall design: Examination of ZMEL1 cells in vitro versus ~21 days in vivo in the zebrafish
Microenvironment-derived factors driving metastatic plasticity in melanoma.
No sample metadata fieldsView Samples
Mitochondrial biogenesis is regulated by signaling pathways sensitive to extracellular conditions and to the internal environment of the cell. We found that deletion of protein phosphatase 2A (PP2A) or of protein phosphatase 6 (PP6) diminishes the nuclear transcriptional response associated with mtDNA damage. Overall design: Six samples were analyzed to determine message RNA levels.
Deletion of conserved protein phosphatases reverses defects associated with mitochondrial DNA damage in Saccharomyces cerevisiae.
Specimen part, SubjectView Samples
Sympathetic neurons of SCG (Superior Cervical Ganglia) send axonal projections either along the external carotid arteries to innervate the salivary glands, or along the internal carotid arteries to the lacrimal and pineal glands, the eye, blood vessels and skin of the head, and the mucosa of the oral and nasal cavities. Previous studies using Wnt1Cre and R26R have defined the neural crest and mesodermal origins of vascular smooth muscle in the heart outflow tract and great vessels, although not specifically of the segments that are relevant for the projections of the SCG neurons. The third pharyngeal arch arteries are lined by neural crest-derived smooth muscle, and consequently, their derivatives, including the entirety of the external carotid arteries and only the base of the internal carotid arteries, also have a neural crest origin. In contrast, the dorsal aortae are lined by smooth muscle that is mesodermal in origin, and as a result, the internal carotid arteries from just above their origination from the common carotid arteries have a mesoderm-derived smooth muscle layer. To address the possibility that guidance cues for SCG neurons are selectively expressed by the external carotid vs. the internal carotid arteries, we isolated these segments of the vasculature from mouse embryos at E13.5 and extracted RNA to screen microarrays for differentially expressed genes.
Endothelins are vascular-derived axonal guidance cues for developing sympathetic neurons.
No sample metadata fieldsView Samples
Basic helix-loop-helix (bHLH) proneural transcription factors (TFs) Ascl1 and Neurog2 are integral to the development of the nervous system. Here, we investigated the molecular mechanisms by which Ascl1 and Neurog2 control the acquisition of generic neuronal fate and impose neuronal subtype identity. Using direct neuronal programming of embryonic stem cells, we found that Ascl1 and Neurog2 regulate distinct targets by binding to largely different sets of sites. Their divergent binding pattern is not determined by the previous chromatin state but distinguished by specific E-box enrichments which reflect the DNA sequence preference of the bHLH domain. The divergent Ascl1 and Neurog2 binding patterns result in distinct chromatin accessibility and enhancer activity landscapes that shape the binding and activity of downstream TFs during neuronal specification. Our findings suggest that proneural factors contribute to neuronal diversity by differentially altering the chromatin landscapes that shape the binding of neuronally expressed TFs. Overall design: Single-cell RNA-seq was used to characterize gene expression in mixed populations of mES cells containing induced expression of either Ascl1 or Neurog2.
Proneural factors Ascl1 and Neurog2 contribute to neuronal subtype identities by establishing distinct chromatin landscapes.
Specimen part, Treatment, SubjectView Samples
Human umbilical cord Whartons jelly stem cells (WHJSC) are gaining attention as a possible clinical source of mesenchymal stem cells for use in cell therapy and tissue engineering due to their high accessibility, expansion potential and plasticity. However, the cell viability changes that are associated to sequential cell passage of these cells are not known. In this analysis, we have identified the gene expression changes that are associated to cell passage in WHJSC.
Evaluation of the cell viability of human Wharton's jelly stem cells for use in cell therapy.
Specimen partView Samples
Compare the behaviour of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow. The effect of culture conditions on the behaviour of MSC was also characterised by isolating MSC and then culturing the cells for 96h in MAPC growth conditions
Validation of COL11A1/procollagen 11A1 expression in TGF-β1-activated immortalised human mesenchymal cells and in stromal cells of human colon adenocarcinoma.
Age, Specimen partView Samples
X-chromosome inactivation (XCI) provides a dosage compensation mechanism where, in each female cell, one of the two X chromosomes is randomly silenced. However, some genes on the inactive X chromosome and outside the pseudoautosomal regions escape from XCI and are expressed from both alleles (escapees). We investigated XCI at single-cell resolution combining deep single cellRNA sequencing with whole-genome sequencing to examine allelic-specific expression in 935 primary fibroblast and 48 lymphoblastoid single cells from five female individuals. In this framework we integrated an original method to identify and exclude doublets of cells. In fibroblast cells, we have identified 55 genes as escapees including five novel escapee genes. Moreover, we observed that all genes exhibit a variable propensity to escape XCI in each cell and cell type and that each cell displays a distinct expression profile of the escapee genes. A metric, the Inactivation Score—defined as the mean of the allelic expression profiles of the escapees per cell—enables us to discover a heterogeneous and continuous degree of cellular XCI with extremes represented by “inactive” cells, i.e., cells exclusively expressing the escaping genes from the active X chromosome and “escaping” cells expressing the escapees from both alleles. We found that this effect is associated with cell-cycle phases and, independently, with the XIST expression level, which is higher in the quiescent phase (G0). Single-cell allele-specific expression is a powerful tool to identify novel escapees in different tissues and provide evidence of an unexpected cellular heterogeneity of XCI. Overall design: Single-cell RNA seq study on 935 human fibroblasts and 48 lymphoblastoid cells from 5 female individuals, in order to investigate the X chromosome nactivation mechanism on a single cell level and to identify escapee genes
Single cell transcriptome in aneuploidies reveals mechanisms of gene dosage imbalance.
Specimen part, SubjectView Samples
Direct programming via the overexpression of transcription factors (TFs) aims to control cell fate at a precision that will be instrumental for clinical applications. However, direct programming of terminal fates remains an obscure process. Taking advantage of the rapid and uniquely efficient programming of spinal motor neurons by overexpression of Ngn2, Isl1 and Lhx3, we have characterized gene expression, chromatin and transcription factor binding time-course dynamics during complete motor neuron programming. Our studies point to a surprisingly dynamic programming process. Promoter chromatin and expression analysis reveals at least three distinct phases of gene activation, while programming factor binding shifts from one set of targets to another, controlling regulatory region activity and gene expression. Furthermore, our evidence suggest that the enhancers and genes activated in the final stage of motor neuron processing are dependent on the combined activities of Isl1 and Lhx3 factors with Ebf and Onecut TFs that are themselves activated midway through the programming process. Our results suggest an unexpected multi-stage model of motor neuron programming in which the programming TFs require activation of a set of intermediate regulators before they complete the programming process. Overall design: Gene expression was characterized by single-cell RNA-seq during the direct programming of ES cells into motor neurons using over-expression of Ngn2-Isl1-Lhx3 programming factors.
A Multi-step Transcriptional and Chromatin State Cascade Underlies Motor Neuron Programming from Embryonic Stem Cells.
Specimen part, Cell line, Treatment, SubjectView Samples
This SuperSeries is composed of the SubSeries listed below.
Integrative analysis of DNA copy number, DNA methylation and gene expression in multiple myeloma reveals alterations related to relapse.
Sex, Age, Specimen partView Samples