Ena-DATASET-WTCHG-06-03-2019-11:18:49:142-220 - samples
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ABSTRACT:
PROVIDER: EGAD00001004823 | EGA |
REPOSITORIES: EGA
Similar Datasets
Project description:This dataset contains 27 mapped bam files. The samples were generated with 3 different protocols for deriving pancreatic progenitors from hPSC. Three parallel differentiations were performed, all done in a hPSC NKX6.1-GFP reporter line. For each protocol there are three cellular populations: total (presort), GFP+ and GFP- . In summary: 3 differentiations x 3 protocols x 3 cellular populations. We prepared ATAC-seq libraries for the 27 samples, and sequenced them on Illumina HiSeq4000.
| EGAD00001004824 | EGA
Project description:We generated human induced pluripotent stem cell (iPSC) lines with a GFP reporter inserted in the endogenous NKX6.1 locus. Characterisation of the reporter lines demonstrated faithful GFP labelling of NKX6.1 expression during pancreas and motor neuron differentiation. We performed three independent in vitro differentiations towards the pancreatic endocrine lineage. We FACS-purified GFP positive and negative cells from stage 7 cultures, and generated Smart-Seq2 RNA-sequencing libraries for the pre-sorted cells, as well as the two GFP-sorted cell populations. Gene expression profiling by RNA-sequencing reveals that the NKX6.1-positive population closely resembles mature human beta cells and the functional evaluation of purified populations shows that the glucose-responsive beta-like cells are enriched within the NKX6.1-positive population. These reporter lines provide a valuable resource to the scientific community for the derivation of functional relevant pancreas and neuronal cell subtypes.
| EGAD00001003911 | EGA
Project description:Human pluripotent stem cells (hPSCs) provide both powerful models for studying cellular differentiations, and unlimited sources of cells for regenerative medicine. However, a comprehensive single cell level differentiation roadmap for hPSCs has not been achieved yet. Here, we used high throughput single-cell RNA-sequencing (scRNA-seq) method, based on optimized microfluidic circuits, to profile early differentiation lineages in the human embryoid body (EB) system. We presented a cellular landscape for hPSC early differentiations covering different cellular lineages, including neural cell, muscle cell, endothelial cell, stromal cell, liver cell, and epithelial cell. Through pseudotemporal analysis, we constructed the differentiation trajectories of these progenitor cells and revealed the gene expression dynamics in the process of differentiation. We reset Primed H9 into Naïve-like H9 and studied cell state transition process via scRNA-seq. We found that mesendoderm genes are enriched in Naïve-like H9. Functionally, Naïve-like H9 showed better potency for differentiation into the hematopoietic lineages. We constructed the differentiation landscape of hPSC early differentiation by scRNA-seq analysis. We offer new insights into molecular pathways of early embryonic lineages that can be harnessed for optimization of differentiation protocols. (Pipeline for C1 data demultiplex : https://github.com/bioeauty/sccpipe)
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Project description:Lung and hindgut directed differentiations with multiple protocols and time points
2020-01-13 | GSE128922 | GEO
Project description:We generated human induced pluripotent stem cell (iPSC) lines with a GFP reporter inserted in the endogenous NKX6.1 locus. Characterisation of the reporter lines demonstrated faithful GFP labelling of NKX6.1 expression during pancreas and motor neuron differentiation. We performed three independent in vitro differentiations towards the pancreatic endocrine lineage. We FACS-purified GFP positive and negative cells from stage 7 cultures, and generated Smart-Seq2 RNA-sequencing libraries for the pre-sorted cells, as well as the two GFP-sorted cell populations. Gene expression profiling by RNA-sequencing reveals that the NKX6.1-positive population closely resembles mature human beta cells and the functional evaluation of purified populations shows that the glucose-responsive beta-like cells are enriched within the NKX6.1-positive population. These reporter lines provide a valuable resource to the scientific community for the derivation of functional relevant pancreas and neuronal cell subtypes.
| EGAS00001002802 | EGA
Project description:Neurons derived from human pluripotent stem cells (hPSCs) are a remarkable tool for modeling human neural development and diseases. However, it remains largely unknown whether the hPSC-derived neurons can be functionally coupled with their target tissues in vitro, which is essential for understanding inter-cellular physiology and further translational studies. Here, we demonstrate that hPSC-derived sympathetic neurons can be obtained from hPSCs and that the resulting neurons form physical and functional connections with cardiac muscle cells. By use of multiple hPSC reporter lines, we recapitulated human autonomic neuron development in vitro, and successfully isolated PHOX2B::eGFP+ neurons exhibiting sympathetic marker expression, electrophysiological properties, and norepinephrine secretion. With pharmacological and optogenetic manipulations, the PHOX2B::eGFP+ neurons controlled the beating rates of cardiomyocytes, and their physical interaction led to neuronal maturation. Our study lays a foundation for the specification of human sympathetic neurons and for the hPSC-based neuronal control of end organs in a dish. Using the four genetic reporter systems (OCT4::eGFP, SOX10::eGFP, ASCL1::eGFP, and PHOX2B::eGFP reporter hESC lines), we were able to purify discrete cell populations at four differentiation stages, recapitulating the sympathoadrenal differentiation process in vitro with purified and defined populations in four specific differentiation stages. We performed transcriptome analysis of OCT4::eGFP+ cells (3 biological replicates, representing undifferentiated hESCs), SOX10::eGFP+ cells (3 biological replicates, multi-potent neural crest), ASCL1::eGFP+ cells (3 biological replicates, putative sympathoadrenal progenitors), and PHOX2B::eGFP+ cells (2 biological replicates, putative sympathetic neuronal precursors).
2016-07-01 | E-GEOD-80689 | biostudies-arrayexpress
Project description:Abstract: Human pluripotent stem cells (hPSCs) can be directed to differentiate into skeletal muscle progenitor cells (SMPCs). However, the myogenic potential of hPSC-SMPCs compared to human fetal or adult satellite cells (Scs) remains unclear. This study demonstrates hPSC-SMPCs derived by commonly used protocols are functionally less mature than freshly-isolated human fetal or adult Scs. We utilized RNA-SEQ of human fetal Scs to identify differentially expressed genes including NGFR, ERBB3, which enriched for MYOD+ or PAX7+ cells. Furthermore, inhibition of TGFβ (TGFβi) improved hPSC-muscle maturation in three independent lines as measured by TEM and expression of myosins. Engraftment of CRISPR/Cas9-corrected Duchenne Muscular Dystrophy (DMD) hiPSC-SMPC subpopulations treated with TGFβi in vivo, increased dystrophin-positive fibers to levels of engrafted cultured fetal Scs in mdx mouse models of DMD. This work provides the first characterization of the developmental status of hPSC-SMPCs and identifies candidates to enable robust myogenic activity in vitro and in vivo.
2016-09-27 | GSE87365 | GEO
Project description:Next-generation sequencing (NGS) has significantly facilitated the analysis of the gene profile and elucidated the molecular mechanisms important for specific cell lineage differentiated from human pluripotent stem cells (hPSCs). Here we report the application of RNA-sequencing technology for transcriptome profile of primary human brain microvascular endothelial cells (BMECs) and hPSC-derived BMECs, and comparison of transcriptomes among cells, including hPSCs, mesodermal progenitors, ectodermal progenitors, endodermal progenitors, hBMECs and hPSC-derived BMECs from hPSCs. Four different BMEC samples differentiated from hPSCs by two different methods and hBMECs were performed in IIIumina HiSeq2500. The resulting sequence reads (about 20 million reads per sample) were mapped to human genome (hg19) using HISAT, and the RefSeq transcript levels (FPKMs) were quantified using the python script rpkmforgenes.py. We next analyzed the expression of a subset of genes that regulate key BBB attributes, including tight junctions and molecular transporters. The gene set comprises 20 tight junction-related genes and an unbiased list of all 25 CLDN genes, all 407 solute carrier (SLC) transporters, and all 53 ATP-binding cassette (ABC) transporters regardless of prior knowledge of BBB association. Primary human BMECs expressed 234 of these genes. BMECs differentiated from hPSCs via the defined method expressed many of these same genes (206 of 234 (88%)) as did BMECs differentiated via the UM method (208 of 234 (89%), indicating a close similarity between hPSC-derived BMECs and primary human BMECs with respect to transcripts having likely relevance to BBB function. RNA sequencing was used to compare global gene expression profiles in the hPSC-derived BMECs with BMECs generated from our previously reported co-differentiation system and primary human BMECs. As expected, hPSC-derived BMECs from three independent differentiations clustered closely and were similar to those generated from the undefined UM platform. Moreover, the hPSC-derived BMECs clustered with primary human BMECs and were distinct from undifferentiated hPSCs and hPSC-derived ectoderm, endoderm and mesodermThis study provides detailed transcriptome comparison between hBMECs and hPSC-derived BMECs and unveils important genes related BMEC phenotypes.
2017-10-10 | GSE97575 | GEO
Project description:Despite the progress in safety and efficacy of cell therapy with pluripotent stem cells (PSCs), the presence of residual undifferentiated stem cells or proliferating neural progenitor cells (NPCs) with rostral identity has remained a major challenge. Here we reported the generation of an LMX1A knock-in GFP reporter human embryonic stem cell (hESC) line that marks the early dopaminergic progenitors during neural differentiation. Purified GFP positive cells in vitro exhibited expression of mRNA and proteins that characterized and matched the midbrain dopaminergic identity. Further proteomic analysis of enriched LMX1A+ cells identified several membrane associated proteins including CNTN2, enabling prospective isolation of LMX1A+ progenitor cells. Transplantation of hPSC-derived purified CNTN2+ progenitors enhanced dopamine release from transplanted cells in the host brain and alleviated Parkinson’s disease symptoms in animal models. Our study establishes an efficient approach for purification of large numbers of hPSC-derived dopaminergic progenitors for therapeutic applications.
2018-06-06 | PXD007837 | Pride
Project description:The Ventx genes are a family of non-clustered homeobox transcription factors that confer a ventral phenotype on mesodermal cells in the Xenopus laevis embryo. Ventx genes are conserved in vertebrates, although they have been lost in rodents. We evaluated the expression of VENTX during differentiation of a human pluripotent stem cell (hPSC) line in which a GFP reporter marked the expression of VENTX (VENTXGFP/w), discovering that VENTX was expressed in a subset of early primordial germ cell-like cells (PGCLCs). We generated a dual reporter hPSC line in which mCHERRY marked the pluripotency gene OCT4 (VENTXGFP/wOCT42AmCHERRY/w), and further modified this to also create a VENTX knock out line by deleting the VENTX homeodomain (VENTXGFP/-OCT42AmCHERRY/w). Transcriptional profiling experiments revealed two populations of PGCLCs, those that co expressed VENTX and OCT4 (GFP+mCHERRY+) and those that were VENTX-OCT4+ (GFP-mCHERRY+). Our analyses showed that VENTX+ and VENTX- PGCLCs clustered together, and also failed to reveal transcriptional consequences of VENTX deletion on commitment to PGCLCs. Evaluation of public databases confirmed expression of VENTX in populations of PGCLCs derived by others and in primary human embryonic germ cell samples. These results identify VENTX as a new PGC-associated gene in humans, however it does not seem to be required for initial human PGC commitment.
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