Derivation of endothelial cells from human embryonic stem cells in fully defined medium enables identification of lysophosphatidic acid and platelet activating factor as regulators of eNOS localization

Magdaline Costa, Koula Sourris, Sue Mei Lim, Qing Cissy Yu, Claire Elizabeth Hirst, Helena Cecilia Parkington, Vanta Joanna Jameson-Jokubaitis, Anthony Edwin Dear, Hong Bin Liu, Suzanne Jeanine Micallef, Katerina Koutsis, Andrew George Elefanty, Edouard Stanley

Research output: Contribution to journalArticleResearchpeer-review

Abstract

The limited availability of human vascular endothelial cells (ECs) hampers research into EC function whilst the lack of precisely defined culture conditions for this cell type presents problems for addressing basic questions surrounding EC physiology. We aimed to generate endothelial progenitors from human pluripotent stem cells to facilitate the study of human EC physiology, using a defined serum-free protocol. Human embryonic stem cells (hESC-ECs) differentiated under serum-free conditions generated CD34(+)KDR(+) endothelial progenitor cells after 6days that could be further expanded in the presence of vascular endothelial growth factor (VEGF). The resultant EC population expressed CD31 and TIE2/TEK, took up acetylated low-density lipoprotein (LDL) and up-regulated expression of ICAM-1, PAI-1 and ET-1 following treatment with TNFalpha. Immunofluorescence studies indicated that a key mediator of vascular tone, endothelial nitric oxide synthase (eNOS), was localised to a perinuclear compartment of hESC-ECs, in contrast with the pan-cellular distribution of this enzyme within human umbilical vein ECs (HUVECs). Further investigation revealed that that the serum-associated lipids, lysophosphatidic acid (LPA) and platelet activating factor (PAF), were the key molecules that affected eNOS localisation in hESC-ECs cultures. These studies illustrate the feasibility of EC generation from hESCs and the utility of these cells for investigating environmental cues that impact on EC phenotype. We have demonstrated a hitherto unrecognized role for LPA and PAF in the regulation of eNOS subcellular localization.
Original languageEnglish
Pages (from-to)103 - 117
Number of pages15
JournalStem Cell Research
Volume10
Issue number1
DOIs
Publication statusPublished - 2013

Cite this

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title = "Derivation of endothelial cells from human embryonic stem cells in fully defined medium enables identification of lysophosphatidic acid and platelet activating factor as regulators of eNOS localization",
abstract = "The limited availability of human vascular endothelial cells (ECs) hampers research into EC function whilst the lack of precisely defined culture conditions for this cell type presents problems for addressing basic questions surrounding EC physiology. We aimed to generate endothelial progenitors from human pluripotent stem cells to facilitate the study of human EC physiology, using a defined serum-free protocol. Human embryonic stem cells (hESC-ECs) differentiated under serum-free conditions generated CD34(+)KDR(+) endothelial progenitor cells after 6days that could be further expanded in the presence of vascular endothelial growth factor (VEGF). The resultant EC population expressed CD31 and TIE2/TEK, took up acetylated low-density lipoprotein (LDL) and up-regulated expression of ICAM-1, PAI-1 and ET-1 following treatment with TNFalpha. Immunofluorescence studies indicated that a key mediator of vascular tone, endothelial nitric oxide synthase (eNOS), was localised to a perinuclear compartment of hESC-ECs, in contrast with the pan-cellular distribution of this enzyme within human umbilical vein ECs (HUVECs). Further investigation revealed that that the serum-associated lipids, lysophosphatidic acid (LPA) and platelet activating factor (PAF), were the key molecules that affected eNOS localisation in hESC-ECs cultures. These studies illustrate the feasibility of EC generation from hESCs and the utility of these cells for investigating environmental cues that impact on EC phenotype. We have demonstrated a hitherto unrecognized role for LPA and PAF in the regulation of eNOS subcellular localization.",
author = "Magdaline Costa and Koula Sourris and Lim, {Sue Mei} and Yu, {Qing Cissy} and Hirst, {Claire Elizabeth} and Parkington, {Helena Cecilia} and Jameson-Jokubaitis, {Vanta Joanna} and Dear, {Anthony Edwin} and Liu, {Hong Bin} and Micallef, {Suzanne Jeanine} and Katerina Koutsis and Elefanty, {Andrew George} and Edouard Stanley",
year = "2013",
doi = "10.1016/j.scr.2012.10.003",
language = "English",
volume = "10",
pages = "103 -- 117",
journal = "Stem Cell Research",
issn = "1873-5061",
publisher = "Elsevier",
number = "1",

}

Derivation of endothelial cells from human embryonic stem cells in fully defined medium enables identification of lysophosphatidic acid and platelet activating factor as regulators of eNOS localization. / Costa, Magdaline; Sourris, Koula; Lim, Sue Mei; Yu, Qing Cissy; Hirst, Claire Elizabeth; Parkington, Helena Cecilia; Jameson-Jokubaitis, Vanta Joanna; Dear, Anthony Edwin; Liu, Hong Bin; Micallef, Suzanne Jeanine; Koutsis, Katerina; Elefanty, Andrew George; Stanley, Edouard.

In: Stem Cell Research, Vol. 10, No. 1, 2013, p. 103 - 117.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Derivation of endothelial cells from human embryonic stem cells in fully defined medium enables identification of lysophosphatidic acid and platelet activating factor as regulators of eNOS localization

AU - Costa, Magdaline

AU - Sourris, Koula

AU - Lim, Sue Mei

AU - Yu, Qing Cissy

AU - Hirst, Claire Elizabeth

AU - Parkington, Helena Cecilia

AU - Jameson-Jokubaitis, Vanta Joanna

AU - Dear, Anthony Edwin

AU - Liu, Hong Bin

AU - Micallef, Suzanne Jeanine

AU - Koutsis, Katerina

AU - Elefanty, Andrew George

AU - Stanley, Edouard

PY - 2013

Y1 - 2013

N2 - The limited availability of human vascular endothelial cells (ECs) hampers research into EC function whilst the lack of precisely defined culture conditions for this cell type presents problems for addressing basic questions surrounding EC physiology. We aimed to generate endothelial progenitors from human pluripotent stem cells to facilitate the study of human EC physiology, using a defined serum-free protocol. Human embryonic stem cells (hESC-ECs) differentiated under serum-free conditions generated CD34(+)KDR(+) endothelial progenitor cells after 6days that could be further expanded in the presence of vascular endothelial growth factor (VEGF). The resultant EC population expressed CD31 and TIE2/TEK, took up acetylated low-density lipoprotein (LDL) and up-regulated expression of ICAM-1, PAI-1 and ET-1 following treatment with TNFalpha. Immunofluorescence studies indicated that a key mediator of vascular tone, endothelial nitric oxide synthase (eNOS), was localised to a perinuclear compartment of hESC-ECs, in contrast with the pan-cellular distribution of this enzyme within human umbilical vein ECs (HUVECs). Further investigation revealed that that the serum-associated lipids, lysophosphatidic acid (LPA) and platelet activating factor (PAF), were the key molecules that affected eNOS localisation in hESC-ECs cultures. These studies illustrate the feasibility of EC generation from hESCs and the utility of these cells for investigating environmental cues that impact on EC phenotype. We have demonstrated a hitherto unrecognized role for LPA and PAF in the regulation of eNOS subcellular localization.

AB - The limited availability of human vascular endothelial cells (ECs) hampers research into EC function whilst the lack of precisely defined culture conditions for this cell type presents problems for addressing basic questions surrounding EC physiology. We aimed to generate endothelial progenitors from human pluripotent stem cells to facilitate the study of human EC physiology, using a defined serum-free protocol. Human embryonic stem cells (hESC-ECs) differentiated under serum-free conditions generated CD34(+)KDR(+) endothelial progenitor cells after 6days that could be further expanded in the presence of vascular endothelial growth factor (VEGF). The resultant EC population expressed CD31 and TIE2/TEK, took up acetylated low-density lipoprotein (LDL) and up-regulated expression of ICAM-1, PAI-1 and ET-1 following treatment with TNFalpha. Immunofluorescence studies indicated that a key mediator of vascular tone, endothelial nitric oxide synthase (eNOS), was localised to a perinuclear compartment of hESC-ECs, in contrast with the pan-cellular distribution of this enzyme within human umbilical vein ECs (HUVECs). Further investigation revealed that that the serum-associated lipids, lysophosphatidic acid (LPA) and platelet activating factor (PAF), were the key molecules that affected eNOS localisation in hESC-ECs cultures. These studies illustrate the feasibility of EC generation from hESCs and the utility of these cells for investigating environmental cues that impact on EC phenotype. We have demonstrated a hitherto unrecognized role for LPA and PAF in the regulation of eNOS subcellular localization.

UR - http://www.sciencedirect.com/science/article/pii/S1873506112001055

U2 - 10.1016/j.scr.2012.10.003

DO - 10.1016/j.scr.2012.10.003

M3 - Article

VL - 10

SP - 103

EP - 117

JO - Stem Cell Research

JF - Stem Cell Research

SN - 1873-5061

IS - 1

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