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Mouse embryonic and induced pluripotent stem cells can form definitive endoderm despite differences in imprinted genes

ABSTRACT: The directed differentiation of induced pluripotent stem (iPS) and embryonic stem (ES) cells into definitive endoderm (DE) would allow the derivation of otherwise inaccessible progenitors for endodermal tissues. However, a global comparison of the relative equivalency of DE derived from iPS and ES populations has not been performed. Recent reports of molecular differences between iPS and ES cells have raised uncertainty as to whether iPS cells could generate autologous endodermal lineages in vitro. Here, we have shown that both mouse iPS and parental ES cells exhibited highly similar in vitro capacity to undergo directed differentiation into DE progenitors. With few exceptions, both cell types displayed similar surges in gene expression of specific master transcriptional regulators and global transcriptomes that define the developmental milestones of DE differentiation. Microarray analysis showed considerable overlap between the genetic programs of DE derived from ES/iPS cells in vitro and authentic DE from mouse embryos in vivo. Intriguingly, iPS cells exhibited aberrant silencing of imprinted genes known to participate in endoderm differentiation, yet retained a robust ability to differentiate into DE. Our results show that, despite some molecular differences, iPS cells can be efficiently differentiated into DE precursors, reinforcing their potential for development of cell-based therapies for diseased endodermal-derived tissues.

ORGANISM(S): Mus musculus

PROVIDER: GSE27087 | GEO | 2011/03/31

SECONDARY ACCESSION(S): PRJNA137467

REPOSITORIES: GEO

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Mouse embryonic and induced pluripotent stem cells can form definitive endoderm despite differences in imprinted genes

Project description:The directed differentiation of induced pluripotent stem (iPS) and embryonic stem (ES) cells into definitive endoderm (DE) would allow the derivation of otherwise inaccessible progenitors for endodermal tissues. However, a global comparison of the relative equivalency of DE derived from iPS and ES populations has not been performed. Recent reports of molecular differences between iPS and ES cells have raised uncertainty as to whether iPS cells could generate autologous endodermal lineages in vitro. Here, we have shown that both mouse iPS and parental ES cells exhibited highly similar in vitro capacity to undergo directed differentiation into DE progenitors. With few exceptions, both cell types displayed similar surges in gene expression of specific master transcriptional regulators and global transcriptomes that define the developmental milestones of DE differentiation. Microarray analysis showed considerable overlap between the genetic programs of DE derived from ES/iPS cells in vitro and authentic DE from mouse embryos in vivo. Intriguingly, iPS cells exhibited aberrant silencing of imprinted genes known to participate in endoderm differentiation, yet retained a robust ability to differentiate into DE. Our results show that, despite some molecular differences, iPS cells can be efficiently differentiated into DE precursors, reinforcing their potential for development of cell-based therapies for diseased endodermal-derived tissues. Comparison of mouse ES cells, mouse iPS cells, and E8.25 Mouse embryos; in the undifferentiated state and definitive endoderm differentiated state. ckit+/Sox2dim =definitive endoderm (day 5 of differentiation in vitro); Sox2bright =undifferentiated ES or iPS cells (day 0 of differentiation); ENDM1+/Epcam+/SSlo=foregut endoderm sorted from mouse E8.25 embryos; Epcam+/ENDM1 negative =sorted comparison population from mouse E8.25 embryos.

2011-03-31 | E-GEOD-27087 | biostudies-arrayexpress

Gene expression of human embryonic stem cells during endoderm differentiation

Project description:Using a well-established human definitive endoderm differentiation system, we can analyze the gene expression dynamic of human embryonic stem cells (hESCs) during endoderm differentiation. Samples from hESCs, D2 cells and definitive endoderm cells (DE) were collected for deep sequencing with two replicates for each sample. Differentially expressed genes analysis showed that endodermal genes were induced during endoderm differentiation.

2022-01-21 | GSE168625 | GEO

Generation of organized anterior foregut epithelia from pluripotent stem cells using small molecules

Project description:Anterior foregut endoderm (AFE) gives rise to many tissue types of interest for therapeutic research including the esophagus, salivary glands, lung, thymus, parathyroid and thyroid. Despite its importance, only few reports describe the generation of AFE from pluripotent stem cells (PSCs) by directed differentiation. Here, we describe a novel protocol to derive a subdomain of AFE, identified by expression of Pax9, from PSCs using small molecules and chemically defined conditions. Generation of a reporter PSC line allows isolation and characterization of Pax9+ AFE cells. When transplanted in vivo, Pax9+ AFE can form several distinct types of complex anterior foregut epithelia including mucosal glands and stratified squamous epithelium. Finally, we show that the directed differentiation protocol can be used to generate AFE from DiGeorge Syndrome patient-specific human induced PSCs, thus creating a platform to produce anterior foregut derivatives for therapy and to enable the study of disorders of the AFE. Total RNA obtained from FACS purified from in vitro dervied mouse definitive endoderm, anterior foregut and ES cells. AFE cells were derived from a 129X1/SvJ background, DE cells from 129X1/SvJ x 129S1/SV-+p+Tyr- cKitlSl-J/+ (R1 ES cells) and non reporter ES cells from a 129P2/OlaHsd background.

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NextGen Consortium: Globin Gene Expression in Sickle Cell Genotype-Specific iPS Cells

Project description:<p> <ol> <li>Implement an efficient, highly reproducible and 'scalable' system for the production of large numbers of sickle cell anemia-specific iPS cells (iPSC).</li> <li>Derive and characterize a novel, in vitro system for the production of an unlimited supply of erythroid lineage cells from the directed differentiation of 'clinical grade' transgene-free iPS cells; use this system to recapitulate erythroid-lineage ontogeny in vitro with the sequential development of primitive and definitive erythropoiesis, accompanied by the appropriate expression of stage-specific globin genes.</li> <li>Identify developmental gene expression profile differences between erythroid precursors that produce primarily HbF and those that produce primarily HbA or HbS.</li> <li>Determine the effects of the three known HbF major quantitative trait loci (QTL) on globin gene expression in disease-specific iPS cells during in vitro erythropoiesis.</li> <li>Search for novel HbF genetic modifiers associated with markedly elevated HbF levels found in sickle cell anemia patients naturally, or in response to hydroxyurea treatment, by examining gene expression profiles and mRNA sequence of their iPSC-derived erythroid cells.</li> <li>Develop and use a CRISPR-based gene editing platform to study the effect of novel HbF genetic modifiers, explore globin switching, and correct the HbS mutation in sickle iPSC lines.</li> </ol> </p>

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Factors from human embryonic stem cell-derived fibroblast-like cells promote topology-dependent hepatic differentiation in primate embryonic stem cells and human induced pluripotent stem cells

Project description:The future clinical use of embryonic stem cell (ESC)-based hepatocyte replacement therapy depends on the development of an efficient procedure for differentiation of hepatocyes from ESCs. Here we report that a high density of human embryonic stem cell (ESC)-derived fibroblast-like cells (hESdFs) supported the efficient generation of hepatocyte-like cells (HLCs) with functional and mature hepatic phenotypes from primate ESCs and human induced pluripotent stem cells (iPSCs). Molecular and immunocytochemistry analyses revealed that hESdFs caused a rapid loss of pluripotency and induced a sequential endoderm-to-hepatocyte differentiation in the central area of ESC colonies. Knockdown experiments demonstrated that pluripotent stem cells were directed toward endodermal and hepatic lineages by FGF2 and Activin A secreted from hESdFs. Furthermore, we found that the central region of ESC colonies was essential for the hepatic endoderm-specific differentiation, as its removal caused a complete disruption of endodermal differentiation. In conclusion, we describe a novel in vitro differentiation model, and show that hESdF-secreted factors act in concert with regional features of ESC colonies to induce robust hepatic endoderm differentiation in primate pluripotent stem cells. Total RNA were isolated from ORMES6 ESC, differentiated cells at IVDS2 and 3, and cells in the central foci (IVDS2-C) and peripheral (IVDS2-P) area of ESC colonies at IVDS2. Each condition was repeated twice and used ORMES6 ESC as control.

2010-08-20 | E-GEOD-19964 | biostudies-arrayexpress

Dynamic developmental signaling logic underlying lineage bifurcations during human endoderm induction and patterning from pluripotent stem cells [Expression data set]

Project description:The definitive endoderm germ layer is the provenance of multiple internal organs, including the lungs, liver, pancreas and intestines. Molecular events driving initial endoderm germ layer specification and subsequent anterior-posterior patterning of endoderm into distinct organ primordia remain largely cryptic. Through microarray analyses, we captured genome-wide transcriptional dynamics driving successive stages of endoderm development with the intent of identifying novel regulatory genes or diagnostic markers that respectively drive or mark endoderm committment. HES3 human embryonic stem cells (hESCs) were differentiated into highly homogeneous endodermal progenitor populations, and microarray analyses were conducted of six different populations at different tiers of the endodermal lineage hierarchy: undifferentiated hESCs, anterior primitive streak (day 1 of in vitro differentiation), definitive endoderm (day 3) and anterior foregut, posterior foregut or midgut/hindgut patterned endoderm populations (day 7). Additionally, we compared hESCs differentiated using two alternative endoderm induction protocols, serum-based or AFBLy-based differentiation (both day 3 of differentiation).

2013-11-07 | E-GEOD-52158 | biostudies-arrayexpress

Metabolomic Profiling of Human Pluripotent Stem Cell Differentiation into Lung Progenitors

Project description:Metabolism is vital to cellular function and tissue homeostasis during human lung development. In utero, embryonic pluripotent stem cells undergo endodermal differentiation towards a lung progenitor cell fate that can be mimicked in vitro using induced human pluripotent stem cells (hiPSCs) to study genetic mutations. To identify differences between wild type and surfactant protein B (SFTPB)-deficient cell lines during endoderm specification towards lung, we used an untargeted metabolomics approach to evaluate the developmental changes in metabolites. We found that the metabolites most enriched during the differentiation from pluripotent stem cell to lung progenitor cell, regardless of cell line, were sphingomyelins and phosphatidylcholines, two important lipid classes in fetal lung development. The SFTPB mutation had no metabolic impact on early endodermal lung development. The identified metabolite signatures during lung progenitor cell differentiation may be utilized as biomarkers for normal embryonic lung development.

2021-06-29 | ST002055 | MetabolomicsWorkbench

ena-DATASET-WTCHG-10-10-2017-09:37:07:358-646 - samples

Project description:RNA-seq data obtained from directed differentiation of a subset of FiPSCs and BiPSCs cell lines towards islet-like cells. RNA was collected at two key developmental stages: definitive endoderm (DE) and pancreatic progenitors (PP).

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Factors from human embryonic stem cell-derived fibroblast-like cells promote topology-dependent hepatic differentiation in primate embryonic stem cells and human induced pluripotent stem cells

2010-08-20 | GSE19964 | GEO

Human Pluripotent Cell Derived Endothelial Cell Characterization

Project description:Human ES or iPS Cells were differentiated into endothelial cells (ECs) defined by expression of CD31 (PECAM1) and CD144 (VE-Cadherin) on the cell surface. All ES or iPS derived ECs were greater than 90% double positive for these two markers. These in vitro derived ECs were compared to each other and to ECs from primary cell sources; arterial (aortic Ecs), venous (saphenous vein) and lymphatic. ES and iPS cells were differentiated into Ecs using a directed differentiation protocol using sequential addition of growth factors to recapitulate endogenous differentiation (FGF, BMP4, Activin, VEGF). These ECs were grown until 21 days after the start of differentiation. Primary cells were obtained from a commercial vendor and grown to passage 3 to minimize cellular senescence and in vitro artifacts. The RNA was cleaned and DNAse digested on RNeasy columns (Qiagen, Valencia, CA). RNA was amplified, fragmented and labeled using NuGEN Technologies Applause Plus kits and the manufacturer's instructions. Labeled samples were then hybridized to Affymetrix Human Gene 1.0 ST GeneChips. The GeneChips were then washed and stained on Affymetrix 450 Fluidics Stations and scanned on an Affymetrix 3000 Scanner according to standard protocols.

2012-09-01 | E-GEOD-37631 | biostudies-arrayexpress

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