An Engineered Multicellular Stem Cell Niche for the 3D Derivation of Human Myogenic Progenitors from iPSCs
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ABSTRACT:
PROVIDER: PRJNA831685 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18909800.fastq.gz | Fastqsanger.gz | |||
| SRR18909801.fastq.gz | Fastqsanger.gz | |||
| SRR18909802.fastq.gz | Fastqsanger.gz | |||
| SRR18909803.fastq.gz | Fastqsanger.gz | |||
| SRR18909804.fastq.gz | Fastqsanger.gz |
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Project description:An Engineered Multicellular Stem Cell Niche for the 3D Derivation of Human Myogenic Progenitors from iPSCs
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Project description:Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human induced pluripotent stem cell (hiPSC) differentiation, we screened cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of sequential Wnt and FGF pathway induction, these three-component embryoids (iTCEs) are enriched in Pax7 positive embryonic-like myogenic progenitors (eMPs) that can be isolated by flow cytometry. Myogenic differentiation of hiPSCs in iTCEs relies on a specialized structural microenvironment and depends on MAPK, PI3K/AKT, and Notch signaling. After transplantation in a mouse model of Duchenne muscular dystrophy, eMPs repopulate the stem cell niche, reactivate after repeated injury and, compared to adult human myoblasts, display enhanced fusion and lead to stronger muscles. Altogether, we provide a two-week protocol for efficient and scalable suspension-based 3D derivation of Pax7 positive myogenic progenitors from hiPSCs.
2022-05-12 | GSE201424 | GEO
Project description:Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human induced pluripotent stem cell (hiPSC) differentiation, we screened cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of consecutive Wnt and FGF pathway induction, three-component embryoids (iTCEs) containing these cell types yielded between 40-50% human Pax7 positive embryonic-like myogenic progenitors (eMPs). In a secondary screen we identified cell surface markers allowing for flow cytometric isolation of >99% of the Pax7 positive eMP population from iTCEs. Myogenic differentiation of hiPSCs in iTCEs relies on a specialized structural microenvironment and activates the MAPK, PI3K/AKT, and Notch pathways. After transplantation in a mouse model Duchenne muscular dystrophy, eMPs repopulate the stem cell niche, reactivate after repeated injury and, compared to adult human myoblasts, display enhanced fusion and lead to stronger muscles. Altogether, we provide a highly efficient and scalable suspension-based protocol for 3D derivation of Pax7 positive myogenic progenitors from hiPSCs within a two-week time window.
2022-05-13 | GSE202308 | GEO
Project description:Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human-induced pluripotent stem cell (hiPSC) differentiation, we here screened murine cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of sequential Wnt and FGF pathway induction, these three-component embryoids are enriched in Pax7-positive embryonic-like myogenic progenitors that can be isolated by flow cytometry. Myogenic differentiation of hiPSCs in heterotypic embryoids relies on a specialized structural microenvironment and depends on MAPK, PI3K/AKT, and Notch signaling. After transplantation in a mouse model of Duchenne muscular dystrophy, embryonic-like myogenic progenitors repopulate the stem cell niche, reactivate after repeated injury and, compared to adult human myoblasts, display enhanced fusion and lead to increased muscle function. Altogether, we provide a two-week protocol for efficient and scalable suspension-based 3D derivation of Pax7-positive myogenic progenitors from hiPSCs.
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Project description:Genome-wide gene expression analysis of MyoD-infected DMD-specific iPSCs (GM05112-M5.1) on days 0 (untreated), day 3 and day 8 post Dox treatment, human primary myoblasts (undifferentiated and as differentiated myotubes), and undifferentiated iPSCs from healthy donors (iPSCs-1 and iPSCs-2). DMD-specific iPSCs were infected with lentivirus expressing MyoD under the control of Tet-inducible promoter and another lentivirus expressing the transactivator. To initiate myogenic differentiation, iPSCs were treated with 1µg/ml Dox. RNA was isolated 0, 3 and 8 days later and gene expression analysis was performed.
2014-01-09 | E-GEOD-46633 | biostudies-arrayexpress
Project description:Myogenic differentiation of iPSCs has been done by gene–overexpression or directed differentiation. However, viral integration, long-term culture and the presence of unwanted cells are the main obstacles. By using CRISPR/Cas9n, a novel double reporter hESC line was generated for PAX7/MYF5, allowing prospective readout. This strategy allowed pathway screen to define efficient myogenic induction in hESC/iPSCs. Next, surface marker screen allowed identification of CD10 and CD24 for purification of myogenic progenitors. CD10 expression was also confirmed on human satellite cells and muscle progenitors. In vivo studies using transgene/reporter-free hESC/iPSCs further validated myogenic potential of the cells by dystrophin restoration and satellite cell engraftment in NSG-mdx4cv mice. In addition, side-by-side in vitro and in vivo comparison proved superior specificity of CD10/CD24 compared to recently reported markers (ERBB3/NGFR). This study provides new biological insights for myogenic differentiation using a new cell resource and identifies CD10 as a potential myogenic marker.
2018-12-03 | GSE121154 | GEO
Project description:Hypoxia enhances the reprogramming efficiency of human dermal fibroblasts to become induced pluripotent stem cells (iPSCs). Because we showed previously that the hypoxia facilitates the isolation and maintenance of human dental pulp cells (DPCs), we examined here whether it promotes the reprogramming of DPCs to become iPSCs.
2013-09-03 | GSE45872 | GEO
Project description:Hypoxia enhances the reprogramming efficiency of human dermal fibroblasts to become induced pluripotent stem cells (iPSCs). Because we showed previously that the hypoxia facilitates the isolation and maintenance of human dental pulp cells (DPCs), we examined here whether it promotes the reprogramming of DPCs to become iPSCs. To investigate the effect of oxygen concentration on global gene expression, we compared DPCs cultured for 6 days under hypoxia and normoxia.
2013-09-03 | E-GEOD-45872 | biostudies-arrayexpress
Project description:Induced pluripotent stem cells (iPSCs) can be derived from somatic cells by the introduction of the transcription factors Oct4, Sox2, Klf4 and cMyc using various methods. Here, we describe a new approach for the derivation of murine iPSCs using a polycistronic non-viral inducible vector integrated into pseudo attP sites via the C31 integrase-mediated site-specific recombination and subsequent vector excision by Cre recombinase. The pluripotency of the derived iPSCs was proved by in vitro and in vivo tests. The derived transgene-free iPSCs reactivated the endogenous pluripotency genes like e.g. Oct4, Sox2 and Nanog and the global gene expression profiles of iPSCs lines are highly similar to ESCs and distinct from parental murine fibroblasts. We demonstrated the differentiation potential of iPSCs by generation cells of the three germ layers as well as we successfully created germline chimeric mice from transgene-free iPSCs. In this study, we presented an efficient method for the generation of transgene-free iPSCs using dual-recombinase technology. expression data of iPSCs/ESCs/MEFs
2014-03-29 | E-GEOD-51605 | biostudies-arrayexpress
Project description:Two phases of pluripotency, naïve and primed, have been captured in vitro and studied in details1. A third formative phase was recently proposed to exist between naïve and primed phases2. Formative pluripotency entails permissiveness for direct primordial germ cell (PGC) induction and competence for blastocyst chimeras, and is characterized by transcriptional and epigenetic features intermediate of naïve and primed pluripotency. To date, however, stable pluripotent stem cells (PSCs) harboring formative features haven’t been derived from early mammalian embryos. Here we develop a method which enabled the derivation and culture of stable formative-like embryonic stem cells (ESCs) from mouse blastocysts. Formative-like mouse ESCs share molecular features characteristic of early post-implantation epiblasts and are competent for PGC-like cell induction and blastocyst chimera formation. The same culture also supported the derivation of ESCs and transgene-free induced pluripotent stem cells (iPSCs) from horse blastocysts and fibroblasts, respectively. Horse ESCs/iPSCs transcriptionally resembled mouse formative cells, and could also be directly induced into PGC-like cells. Formative-like horse iPSCs could efficiently chimerize horse, mouse, goat, sheep and pig embryos. Stable formative-like PSCs will be invaluable for studying mammalian pluripotency, and our method may be broadly applicable for the derivation of PGC and chimera competent PSCs from other mammalian species.
2020-11-27 | GSE135990 | GEO