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Single-cell RNA-seq of human cardiovascular progenitor cells generated with a chemically defined, xeno-free laminin based differentiation protocol

ABSTRACT: Single-cell RNA-seq of human cardiovascular progenitor cells generated with a chemically defined, xeno-free laminin based differentiation protocol

PROVIDER: PRJNA525863 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR8690402_1.fastq.gz	Fastqsanger.gz
	SRR8690402_2.fastq.gz	Fastqsanger.gz
	SRR8690403_1.fastq.gz	Fastqsanger.gz
	SRR8690403_2.fastq.gz	Fastqsanger.gz
	SRR8690404_1.fastq.gz	Fastqsanger.gz

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Single-cell RNA-seq of human cardiovascular progenitor cells generated with a chemically defined, xeno-free laminin based differentiation protocol

Project description:Regeneration of injured human heart muscle is limited and an unmet clinical need. There are no methods for the reproducible generation of clinical quality stem-cell-derived cardiovascular progenitors (CVPs). We identified laminin-221 (LN-221) as the most likely expressed cardiac laminin. We produced it as human recombinant protein, and showed that LN-221 promotes differentiation of pluripotent hESCs towards cardiomyocyte lineage and downregulates pluripotency and teratoma associated genes. We developed a chemically defined, xeno-free laminin-based differentiation protocol to generate CVPs. Single-cell RNA sequencing of CVPs derived from hESC lines supported reproducibility and identified three main progenitor subpopulations. These CVPs were transplanted into myocardial infarction mice, where heart function was measured by echocardiogram and human heart muscle bundle formation was identified histologically. This method may provide clinical quality cells for use in regenerative cardiology.

2019-03-20 | GSE127955 | GEO

In vivo generation of post-infarct human cardiac muscle by laminin-promoted cardiovascular progenitors [HS1001 differentiation protocol]

Project description:Regeneration of injured human heart muscle is limited and an unmet clinical need. There are no methods for the reproducible generation of clinical quality stem-cell-derived cardiovascular progenitors (CVPs). We identified laminin-221 (LN-221) as the most likely expressed cardiac laminin. We produced it as human recombinant protein, and showed that LN-221 promotes differentiation of pluripotent hESCs towards cardiomyocyte lineage and downregulates pluripotency and teratoma associated genes. We developed a chemically defined, xeno-free laminin-based differentiation protocol to generate CVPs. We show high reproducibility of the differentiation protocol using time-course bulk RNA sequencing developed from different hESC lines. Single-cell RNA sequencing of CVPs derived from hESC lines supported reproducibility and identified three main progenitor subpopulations. These CVPs were transplanted into myocardial infarction mice, where heart function was measured by echocardiogram and human heart muscle bundle formation was identified histologically. This method may provide clinical quality cells for use in regenerative cardiology.

2019-04-26 | GSE100722 | GEO

In vivo generation of post-infarct human cardiac muscle by laminin-promoted cardiovascular progenitors [H1 differentiation protocol]

Project description:Regeneration of injured human heart muscle is limited and an unmet clinical need. There are no methods for the reproducible generation of clinical quality stem-cell-derived cardiovascular progenitors (CVPs). We identified laminin-221 (LN-221) as the most likely expressed cardiac laminin. We produced it as human recombinant protein, and showed that LN-221 promotes differentiation of pluripotent hESCs towards cardiomyocyte lineage and downregulates pluripotency and teratoma associated genes. We developed a chemically defined, xeno-free laminin-based differentiation protocol to generate CVPs. We show high reproducibility of the differentiation protocol using time-course bulk RNA sequencing developed from different hESC lines. Single-cell RNA sequencing of CVPs derived from hESC lines supported reproducibility and identified three main progenitor subpopulations. These CVPs were transplanted into myocardial infarction mice, where heart function was measured by echocardiogram and human heart muscle bundle formation was identified histologically. This method may provide clinical quality cells for use in regenerative cardiology.

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In vivo generation of post-infarct human cardiac muscle by laminin-promoted cardiovascular progenitors

Project description:This SuperSeries is composed of the SubSeries listed below and contains: bulk RNA-seq of two time series of a xeno-free human cardiomyocyte differentiation protocol starting from HS1001 and H1 hECSs until day 94 cardiomyocytes, single cell sequencing of cardiovascular progenitors (H1 and HS1001-derived cardiovascular progenitors at day 9 and 11) and bulk RNA-Seq of H1 hECSs that have been transferred to a LN-221 matrix. Regeneration of injured human heart muscle is limited and an unmet clinical need. There are no methods for the reproducible generation of clinical quality stem-cell-derived cardiovascular progenitors (CVPs). We identified laminin-221 (LN-221) as the most likely expressed cardiac laminin. We produced it as human recombinant protein, and showed that LN-221 promotes differentiation of pluripotent hESCs towards cardiomyocyte lineage and downregulates pluripotency and teratoma associated genes. We developed a chemically defined, xeno-free laminin-based differentiation protocol to generate CVPs. We show high reproducibility of the differentiation protocol using time-course bulk RNA sequencing developed from different hESC lines. Single-cell RNA sequencing of CVPs derived from hESC lines supported reproducibility and identified three main progenitor subpopulations. These CVPs were transplanted into myocardial infarction mice, where heart function was measured by echocardiogram and human heart muscle bundle formation was identified histologically. This method may provide clinical quality cells for use in regenerative cardiology.

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Xeno-free and Chemically Defined Human System for Culturing Human Epidermal Keratinocytes

Project description:Developing an effective xeno-free and defined system for human keratinocyte culture has been one of the fundamental measures in current cellular therapy. For keratinocytes, in particular, such system will not only fulfill clinical safety and quality criteria, it will allow for the management of less severe burns and other skin injuries such as chronic wounds. To date, the expansion of autologous keratinocytes in the clinical settings still relies on 3T3 feeder co-culture system. Here we report a completely xeno-free and defined culture system by using human recombinant laminins as feeder replacement. We have thoroughly characterized the cells cultured in this new system both in vitro and in vivo. We found that laminin-511/-521, and the unanticipated laminin-411/-421, but not -332, -111, -121, -211, or LN-221 support adult human skin keratinocytes and could maintain their self-renewal properties comparable to the 3T3 system. We believe our new culture method should facilitate broader use of cultured epithelial cell products for today’s regenerative medicine.

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Xeno-free and Chemically Defined Human System for Culturing Human Epidermal Keratinocytes

Project description:Xeno-free and Chemically Defined Human System for Culturing Human Epidermal Keratinocytes

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Gene expression profile of chemically induced expandable cardiovascular progenitor cells derived from mouse fibroblasts

Project description:Cardiovascular progenitor cells (CPCs) can be reprogrammed from somatic fibroblasts by combinations of genes, providing a new avenue for cardiac regenerative therapy. Here we developed a strategy to capture chemically induced CPCs (ciCPCs) during the small molecule-induced cardiac reprogramming of mouse fibroblasts and expand these ciCPCs for a long-term in a chemically defined condition. RNA-seq analysis has been used to compare the expression profile of ciCPCs at early and late passages.

2021-07-01 | GSE159081 | GEO

Differentiation of Human Embryonic Stem Cells to Endothelial Progenitor Cells on Laminins in Defined and Xeno-free Systems

Project description:A major hurdle for in vitro culturing of primary endothelial cells (ECs) is that they readily de-differentiate, hampering their use for therapeutic applications. Human embryonic stem cells (hESCs) may provide an unlimited cell source; however, most current protocols deriving endothelial progenitor cells (EPCs) from hESCs use direct differentiation approaches albeit on undefined matrices, yet final yields are insufficient. We developed a method to culture monolayer hESCs on stem-cell niche laminin (LN) LN511 or LN521 matrix. Here, we report a chemically defined, xeno-free protocol for differentiation of hESCs to EPCs using LN521 as the main culture substrate. We could generate ~95% functional EPCs defined as VEGFR2+CD34+CD31+VE-Cadherin+. RNA sequencing analyses of hESCs, EPCs and primary HUVECs showed differentiation-related ECs expression signatures, regarding basement membrane composition, cell-matrix interactions, and changes in endothelial lineage markers. Our results may facilitate production of stable ECs for treatment of vascular diseases and in vitro cell modeling.

2016-09-29 | GSE85953 | GEO

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Project description:In vivo generation of post-infarct human cardiac muscle by laminin-promoted cardiovascular progenitors

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