The long term goal of our studies is to efficiently derive human endothelial cell types from pluripotent stem cells, such as hES and iPS cells, for human vascular regeneration and clinical therapies. To do so, we need to understand the mechanisms that govern human endothelial cell development, but they have been largely undefined. In our Preliminary Studies, we have begun to dissect the signaling hierarchy that modulates the differentiation of human endothelial cells from pluripotent stem cells, and found that some factors involved in this process differ from those required for murine endothelial cell differentiation. Using a hES cell culture system, we found that Indian Hedgehog (Ihh) promoted endothelial cell differentiation, as evidenced by gene/protein expression, morphology and DiI-AcLDL uptake. Furthermore, inhibition of bone morphogenic protein 4 (BMP4) suppressed endothelial cell formation whereas addition of rhBMP4 to cells treated with the hh inhibitor cyclopamine promoted endothelial cell development. Thus, our Preliminary Studies reveal that Ihh promotes human endothelial cell differentiation from hES cells via BMP signaling, providing novel insights applicable to modulating blood vessel formation for human clinical therapies. The proposed studies will build on our Preliminary Data and further dissect the molecular signaling hierarchy that regulates human endothelial cell differentiation. We will also investigate molecular regulators that promote the generation of specialized human endothelial cell types. Finally, we will optimize our hES cell culture system to enable more efficient generation of endothelial cell types from mesodermal progenitors. Our overarching hypotheses are that the steps that lead to human endothelial cell lineage commitment, differentiation and specialization are dissectible and definable in our pluripotent stem cell model system. Furthermore, by understanding the sequence and kinetics of the molecular events needed for these process, we will gain insight into how to more efficiently derive human endothelial cell types for tissue engineering and clinical therapies for prevalent pathologies. The following Specific Aims will test these hypotheses: 1) further define the mechanism(s) by which Ihh and BMP signaling regulate human differentiation of human endothelial cells. 2) investigate the molecular regulation of human endothelial cell specialization;and 3) establish a feeder-free culture system for efficient differentiation and specialization of endothelial cells from human mesodermal progenitors.

Public Health Relevance

We are studying the process of blood vessel formation and trying to understand how human endothelial cells, that form the lining of blood vessels, are differentiated from human embryonic stem cells that are pluripotent. Insights gained from our studies may help to develop strategies for human tissue regeneration and for treating patients with common vascular diseases.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL096360-05
Application #
8248240
Study Section
Special Emphasis Panel (ZRG1-CVS-F (02))
Program Officer
Schramm, Charlene A
Project Start
2009-04-01
Project End
2014-11-30
Budget Start
2012-12-01
Budget End
2014-11-30
Support Year
5
Fiscal Year
2013
Total Cost
$390,541
Indirect Cost
$154,921
Name
Yale University
Department
None
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
Hirschi, Karen K; Li, Song; Roy, Krishnendu (2014) Induced pluripotent stem cells for regenerative medicine. Annu Rev Biomed Eng 16:277-94
Martin, Kathleen A; Hirschi, Karen K (2014) The magic touch: endothelial cells muscle-up adipose. Circ Res 115:752-4
Co?kun, S├╝leyman; Chao, Hsu; Vasavada, Hema et al. (2014) Development of the fetal bone marrow niche and regulation of HSC quiescence and homing ability by emerging osteolineage cells. Cell Rep 9:581-90
Marcelo, Kathrina L; Goldie, Lauren C; Hirschi, Karen K (2013) Regulation of endothelial cell differentiation and specification. Circ Res 112:1272-87
Fang, Jennifer S; Dai, Cuiping; Kurjiaka, David T et al. (2013) Connexin45 regulates endothelial-induced mesenchymal cell differentiation toward a mural cell phenotype. Arterioscler Thromb Vasc Biol 33:362-8
Ek-Vitorin, Jose F; Burt, Janis M (2013) Structural basis for the selective permeability of channels made of communicating junction proteins. Biochim Biophys Acta 1828:51-68
Goldberg, Joshua S; Vadakkan, Tegy J; Hirschi, Karen K et al. (2013) A computational approach to detect gap junction plaques and associate them with cells in fluorescent images. J Histochem Cytochem 61:283-93
Iacobas, Ionela; Vats, Archana; Hirschi, Karen K (2010) Vascular potential of human pluripotent stem cells. Arterioscler Thromb Vasc Biol 30:1110-7
Chao, Hsu; Hirschi, Karen K (2010) Hemato-vascular origins of endothelial progenitor cells? Microvasc Res 79:169-73
Goldberg, Joshua S; Hirschi, Karen K (2009) Diverse roles of the vasculature within the neural stem cell niche. Regen Med 4:879-97

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