Human Pluripotent Cell Derived Endothelial Cell Characterization
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE37631 | GEO | 2012/09/01
SECONDARY ACCESSION(S): PRJNA162411
REPOSITORIES: GEO
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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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Project description:VEGF induces elongation of endothelial cells (ECs) that are derived from wild-type (Foxo1(+/+)) ES cells. VEGF-induced EC elongation is an important process of angiogenesis. ECs derived from Foxo1(-/-) ES cells fail to elongate in response to VEGF. Gene expression profiling of wild-type and Foxo1(-/-) ECs in the presence or absence of VEGF stimulation identifies responsible genes that regulate EC elongation. One wild-type (Foxo1(+/+)) ES cell clone and one Foxo1(-/-) ES cell clone were used. ES cells were cultured on OP9 stromal cell layer in the presence or absence of VEGF (10 ng/ml). After 6.5 days, VE-cadherin+ CD31+ ECs were isolated by FACS and used for total RNA extraction. Three independent experiments were performed for each condition.
2016-02-16 | E-GEOD-76366 | biostudies-arrayexpress
Project description:Adult endothelial cells (ECs) are known to possess organ-specific gene expression, morphology and function, but whether organ-specific EC gene expression is present during human development is not known. Here, we used bulk RNA-sequencing (RNA-seq) to interrogate the developing human intestine, lung, and kidney in order to identify organ-enriched EC-gene signatures. FACS was used to isolate EC (CD31+CD144+, n=13) and non-EC (CD31-CD144-, n=16) populations from these three organs, profiling at least 4 biological replicates for each organ system. The biological specimens profiled were between 11-20 gestational weeks. We also sequenced cultured human umbilical vein endothelial cells (HUVECs) via bulk RNAseq. Computational approaches were used to identify organ-specific EC-enriched gene signatures across human fetal lung, intestine, and kidney ECs.
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Project description:Endothelial differentiation occurs during normal vascular development in the developing embryo. Mouse embryonic stem (ES) cells were used to further define the molecular mechanisms of endothelial differentiation. By flow cytometry a population of VEGF-R2 positive cells was identified as early as 2.5 days after differentiation of ES cells, and a subset of VEGF-R2 + cells, that were CD41+ positive at 3.5 days. A separate population of VEGF-R2+ stem cells expressing the endothelial-specific marker CD144 (VE-cadherin) was also identified at this same time point. Microarray analysis of >45,000 transcripts was performed on RNA obtained from cells expressing VEGF-R2, CD41, and CD144. Experiment Overall Design: We identified four populations of cells; cells expressing VEGF-R2 (day 2.5), CD41 expressing cells (day 3.5), cells expressing CD144 (VE-Cadherin, day 3.5), and cells expressing CD144 (day 6.5). In addition to this, we have also obtained the negative control cells at each time such as VEGF-R2 (day 2.5) negative, CD41 negative (day 3.5), CD144 negative (VE-Cadherin, day 3.5), and negative CD144 (day 6.5). RNA for the microarray experiments were obtained in duplicate from two separately conducted experiments using the murine embryonic stem cells..
2008-10-20 | E-GEOD-10210 | biostudies-arrayexpress
Project description:Endothelial differentiation occurs during normal vascular development in the developing embryo. Mouse embryonic stem (ES) cells were used to further define the molecular mechanisms of endothelial differentiation. By flow cytometry a population of VEGF-R2 positive cells was identified as early as 2.5 days after differentiation of ES cells, and a subset of VEGF-R2 + cells, that were CD41+ positive at 3.5 days. A separate population of VEGF-R2+ stem cells expressing the endothelial-specific marker CD144 (VE-cadherin) was also identified at this same time point. Microarray analysis of >45,000 transcripts was performed on RNA obtained from cells expressing VEGF-R2, CD41, and CD144. Keywords: expression analysis
2008-07-14 | GSE10210 | GEO
Project description:How cells acquire their fate is a fundamental question in both developmental and regenerative biology. Multipotent progenitors undergo gradual cell fate restriction in response to temporal and positional cues from the microenvironment, the nature of which is far from being clear. In the case of the lymphatic system, venous endothelial cells are thought to give rise to lymphatic vessels, through a process of trans-differentiation. Upon expression of a set of transcription factors, venous cells acquire a lymphatic fate, and bud out to generate the lymphatic vasculature. In this work we challenge this view and show that while lymphatic endothelial cells (LECs) do arise in the Cardinal Vein (CV), they do so from a previously uncharacterized pool of multipotent angioblasts. Using lymphatic-specific transgenic zebrafish, in combination with endothelial photoconvertible reporters, and long-term live imaging, we demonstrate that these multipotent angioblasts can generate not only lymphatic, but also arterious, and venous fates. We further reveal that the underlying endoderm serves as a source of Wnt5b, which acts as a lymphatic inductive signal, promoting the angioblast-to-lymphatic transition. Moreover, Wnt5b induced lymphatic specification in human embryonic stem cells- derived vascular progenitors, suggesting that this process is evolutionary conserved. Our results uncover a novel mechanism of lymphatic vessel formation, whereby multipotent angioblasts and not venous endothelial cells give rise to the lymphatic endothelium, and provide the first characterization of their inductive niche. More broadly, our findings highlight the CV as a plastic and heterogeneous structure containing different cell populations, analogous to the hematopoietic niche in the aortic floor. Following Kaede photoconversion of dorsal or ventral halves of the PCV in Tg(fli1:gal4;uasKaede) embryos at 24 hpf, 6 embryos per group were used for FACS isolation of Kaede photconverted (red) ECs.
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Project description:To investigate what kind of gene expression is regulated downstream of Tie1 signaling, we performed RNA-seq analyses on venous and lymphatic endothelial cells (ECs) between homozygous tie1 mutant embryos and their wild-type and heterozygous siblings during secondary sprouting, respectively. Our analyses indicate that Tie1 signaling controls a series of gene expression involved in lymphatic development including EC migration, proliferation, and differentiation.
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Project description:Background: The formation of the lymphatic system has been a subject of controversy for over a century. Although the accepted view is that lymphatic endothelial cells originate exclusively from veins, there have been reports of non-venous derived lymphatics in multiple tissues. Methods: We performed single-nuclear multiome sequencing analyses on 3,801 individual nuclei isolated from Pax3Cre/+;Rosa26tdTomato mouse embryos at E9.5, the temporal window in which lymphatic endothelium initially emerges. Additionally, we performed bulk ATAC-seq analyses following the OmniATAC protocol on FACS-sorted ECs from Pax3Cre/+;Rosa26tdTomato embryos at E13.5. Results: These analyses revealed the existence of angioblasts which arise from paraxial mesoderm and directly differentiate into lymphatic endothelium. Our findings support the notion that lymphatic endothelial cells also have non-venous origins and explain the discrepancies between previous studies.
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