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Wnt Signaling pp 183-196 | Cite as

Directed Endothelial Progenitor Differentiation from Human Pluripotent Stem Cells Via Wnt Activation Under Defined Conditions

  • Xiaoping Bao
  • Xiaojun Lian
  • Sean P. PalecekEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1481)

Abstract

Efficient derivation of endothelial cells and their progenitors from human pluripotent stem cells (hPSCs) can facilitate studies of human vascular development, disease modeling, drug discovery, and cell-based therapy. Here we provide a detailed protocol for directing hPSCs to functional endothelial cells and their progenitors in a completely defined, growth factor- and serum-free system by temporal modulation of Wnt/β-catenin signaling via small molecules. We demonstrate a 10-day, two-stage process that recapitulates endothelial cell development, in which hPSCs first differentiate to endothelial progenitors that then generate functional endothelial cells and smooth muscle cells. Methods to characterize endothelial cell identity and function are also described.

Key words

Human pluripotent stem cells Endothelial progenitors Endothelial cells Wnt signaling Chemically defined Growth factor-free Serum-free Small molecules 

Notes

Acknowledgments

This study was supported by NIH grant R01 EB007534 and NSF grant EFRI 0735903.

References

  1. 1.
    Kaupisch A, Kennedy L, Stelmanis V et al (2012) Derivation of vascular endothelial cells from human embryonic stem cells under GMP-compliant conditions: towards clinical studies in ischaemic disease. J Cardiovasc Transl Res 5:605–617CrossRefPubMedGoogle Scholar
  2. 2.
    Levenberg S, Golub JS, Amit M et al (2002) Endothelial cells derived from human embryonic stem cells. Proc Natl Acad Sci U S A 99:4391–4396CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Van der Meer AD, Orlova VV, ten Dijke P et al (2013) Three-dimensional co-cultures of human endothelial cells and embryonic stem cell-derived pericytes inside a microfluidic device. Lab Chip 13:3562–3568CrossRefPubMedGoogle Scholar
  4. 4.
    Wilson HK, Canfield SG, Shusta EV, Palecek SP (2014) Concise review: tissue-specific microvascular endothelial cells derived from human pluripotent stem cells. Stem Cells 32:3037–3045CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Ashton RS, Keung AJ, Peltier J, Schaffer DV (2011) Progress and prospects for stem cell engineering. Annu Rev Chem Biomol Eng 2:479–502CrossRefPubMedGoogle Scholar
  6. 6.
    Lian X, Selekman J, Bao X et al (2013) A small molecule inhibitor of SRC family kinases promotes simple epithelial differentiation of human pluripotent stem cells. PLoS One 8:e60016CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Lian X, Bao X, Zilberter M et al (2015) Chemically defined, albumin-free human cardiomyocyte generation. Nat Methods 12:595–596CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Murry CE, Keller G (2008) Differentiation of embryonic stem cells to clinically relevant populations: lessons from embryonic development. Cell 132:661–680CrossRefPubMedGoogle Scholar
  9. 9.
    Choi K-D, Yu J, Smuga-Otto K et al (2009) Hematopoietic and endothelial differentiation of human induced pluripotent stem cells. Stem Cells 27:559–567CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Vodyanik MA, Thomson JA, Slukvin II (2006) Leukosialin (CD43) defines hematopoietic progenitors in human embryonic stem cell differentiation cultures. Blood 108:2095–2105CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    James D, Nam H, Seandel M et al (2010) Expansion and maintenance of human embryonic stem cell-derived endothelial cells by TGFbeta inhibition is Id1 dependent. Nat Biotechnol 28:161–166CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Rufaihah AJ, Huang NF, Jamé S et al (2011) Endothelial cells derived from human iPSCS increase capillary density and improve perfusion in a mouse model of peripheral arterial disease. Arterioscler Thromb Vasc Biol 31:e72–e79CrossRefPubMedGoogle Scholar
  13. 13.
    Goldman O, Feraud O, Boyer-Di Ponio J et al (2009) A boost of BMP4 accelerates the commitment of human embryonic stem cells to the endothelial lineage. Stem Cells 27:1750–1759CrossRefPubMedGoogle Scholar
  14. 14.
    Sahara M, Hansson EM, Wernet O et al (2014) Manipulation of a VEGF-Notch signaling circuit drives formation of functional vascular endothelial progenitors from human pluripotent stem cells. Cell Res 24:820–841CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    White MP, Rufaihah AJ, Liu L et al (2013) Limited gene expression variation in human embryonic stem cell and induced pluripotent stem cell-derived endothelial cells. Stem Cells 31:92–103CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Tatsumi R, Suzuki Y, Sumi T et al (2011) Simple and highly efficient method for production of endothelial cells from human embryonic stem cells. Cell Transplant 20:1423–1430CrossRefPubMedGoogle Scholar
  17. 17.
    Wang ZZ, Au P, Chen T et al (2007) Endothelial cells derived from human embryonic stem cells form durable blood vessels in vivo. Nat Biotechnol 25:317–318CrossRefPubMedGoogle Scholar
  18. 18.
    Bao X, Lian X, Dunn KK et al (2015) Chemically-defined albumin-free differentiation of human pluripotent stem cells to endothelial progenitor cells. Stem Cell Res 15:122–129CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Lian X, Bao X, Al-Ahmad A et al (2014) Efficient differentiation of human pluripotent stem cells to endothelial progenitors via small-molecule activation of WNT signaling. Stem Cell Reports 3:804–816CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Lian X, Zhang J, Azarin SM et al (2013) Directed cardiomyocyte differentiation from human pluripotent stem cells by modulating Wnt/β-catenin signaling under fully defined conditions. Nat Protoc 8:162–175CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Department of Chemical & Biological EngineeringUniversity of WisconsinMadisonUSA
  2. 2.Departments of Biomedical Engineering, Biology, and Huck Institutes of the Life ScienceThe Pennsylvania State UniversityUniversity ParkUSA

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