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Directing human embryonic stem cells towards functional endothelial cells easily and without purification

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Tissue Engineering and Regenerative Medicine Aims and scope

Abstract

Hemangioblasts or blood islands only arise in early development thereby the sources to obtain these bi-potential cells are limited. While previous studies have isolated both lineages in vitro through the hemangioblast, derivation efficiency was rather low due to cellular damage attributed by enzyme usage and fluorescent activated cell sorting (FACS). This study focused on avoiding the use of damaging factors in the derivation of endothelial cells (ECs). Single cell H9-human embryonic stem cells (hESCs) were obtained by using a mild dissociation protocol then human embryoid body (hEB) formation was performed under hemangioblast differentiation conditions. The hEBs were subjected to a two-stage cytokine treatment procedure. Subsequent culture of the adhesive cells in day 4 hEBs gave arise to a seemingly pure population of ECs. The hESC-derived ECs were characterized by identifying signature endothelial gene and protein markers as well as testing for in vitro functionality. Furthermore, in vivo functionality was also confirmed by transplanting the cells in hindlimb ischemic murine models. We demonstrate that the genetic change required for EC derivation precedes blast colony formation. Furthermore, cell damage was prevented by abating enzyme usage and FACS, resulting in a high yield of ECs upon adhesion. Under this method, confluent cultures of ECs were obtainable 4 days after hEB formation which is significantly faster than previous protocols.

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References

  1. Kennedy M, Firpo M, Choi K, Wall C, Robertson S, Kabrun N, et al. A common precursor for primitive erythropoiesis and definitive haematopoiesis. Nature 1997;386:488–493.

    Article  CAS  PubMed  Google Scholar 

  2. Robertson S, Kennedy M, Keller G. Hematopoietic commitment during embryogenesis. Ann N Y Acad Sci 1999;872:9–15. discussion 15-16.

    Article  CAS  PubMed  Google Scholar 

  3. Choi K. The hemangioblast: a common progenitor of hematopoietic and endothelial cells. J Hematother Stem Cell Res 2002;11:91–101.

    Article  PubMed  Google Scholar 

  4. Forrai A, Robb L. The hemangioblast—between blood and vessels. Cell Cycle 2003;2:86–90.

    Article  CAS  PubMed  Google Scholar 

  5. Bollerot K, Pouget C, Jaffredo T. The embryonic origins of hematopoietic stem cells: a tale of hemangioblast and hemogenic endothelium. APMIS 2005;113:790–803.

    Article  PubMed  Google Scholar 

  6. Jaffredo T, Bollerot K, Sugiyama D, Gautier R, Drevon C. Tracing the hemangioblast during embryogenesis: developmental relationships between endothelial and hematopoietic cells. Int J Dev Biol 2005;49:269–277.

    Article  CAS  PubMed  Google Scholar 

  7. Sumpio BE, Riley JT, Dardik A. Cells in focus: endothelial cell. Int J Biochem Cell Biol 2002;34:1508–1512.

    Article  CAS  PubMed  Google Scholar 

  8. Yoder MC. A bipotent mesoderm subset identified via colony-forming assay. Cell Stem Cell 2010;7:643–644.

    Article  CAS  PubMed  Google Scholar 

  9. Cogle CR, Scott EW. The hemangioblast: cradle to clinic. Exp Hematol 2004;32:885–890.

    Article  PubMed  Google Scholar 

  10. Li Z, Han Z, Wu JC. Transplantation of human embryonic stem cell-derived endothelial cells for vascular diseases. J Cell Biochem 2009;106:194–199.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Asahara T, Kawamoto A, Masuda H. Concise review: circulating endothelial progenitor cells for vascular medicine. Stem Cells 2011;29:1650–1655.

    Article  CAS  PubMed  Google Scholar 

  12. Lu SJ, Lee RJ, Napoli C, Oh S, Kimbrel EA, Feng Q. The promise and therapeutic potential of human ES and iPS cells. Stem Cells Int 2011; 2011:959275.

    Article  PubMed  PubMed Central  Google Scholar 

  13. Choi K, Kennedy M, Kazarov A, Papadimitriou JC, Keller G. A common precursor for hematopoietic and endothelial cells. Development 1998;125:725–732.

    CAS  PubMed  Google Scholar 

  14. Levenberg S, Golub JS, Amit M, Itskovitz-Eldor J, Langer R. Endothelial cells derived from human embryonic stem cells. Proc Natl Acad Sci U S A 2002;99:4391–4396.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Huang HL, Hsing HW, Lai TC, Chen YW, Lee TR, Chan HT, et al. Trypsin-induced proteome alteration during cell subculture in mammalian cells. J Biomed Sci 2010;17:36.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Gil CH, Lee JH, Seo J, Park SJ, Park Z, Kim J, et al. Well-defined differentiation of hESC-derived hemangioblasts by embryoid body formation without enzymatic treatment. Biotechnol Lett 2015;37:1315–1322.

    Article  CAS  PubMed  Google Scholar 

  17. Cho SW, Moon SH, Lee SH, Kang SW, Kim J, Lim JM, et al. Improvement of postnatal neovascularization by human embryonic stem cell derived endothelial-like cell transplantation in a mouse model of hindlimb ischemia. Circulation 2007;116:2409–2419.

    Article  CAS  PubMed  Google Scholar 

  18. Kocher AA, Schuster MD, Szabolcs MJ, Takuma S, Burkhoff D, Wang J, et al. Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function. Nat Med 2001;7:430–436.

    Article  CAS  PubMed  Google Scholar 

  19. Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, et al. Embryonic stem cell lines derived from human blastocysts. Science 1998;282:1145–1147.

    Article  CAS  PubMed  Google Scholar 

  20. Kim IG, Hwang MP, Du P, Ko J, Ha CW, Do SH, et al. Bioactive cell-derived matrices combined with polymer mesh scaffold for osteogenesis and bone healing. Biomaterials 2015;50:75–86.

    Article  CAS  PubMed  Google Scholar 

  21. Basak GW, Yasukawa S, Alfaro A, Halligan S, Srivastava AS, Min WP, et al. Human embryonic stem cells hemangioblast express HLA-antigens. J Transl Med 2009;7:27.

    Article  PubMed  PubMed Central  Google Scholar 

  22. Park TS, Zimmerlin L, Zambidis ET. Efficient and simultaneous generation of hematopoietic and vascular progenitors from human induced pluripotent stem cells. Cytometry A 2013;83:114–126.

    Article  PubMed  Google Scholar 

  23. Islam MS, Ni Z, Kaufman DS. Use of human embryonic stem cells to understand hematopoiesis and hematopoietic stem cell niche. Curr Stem Cell Res Ther 2010;5:245–250.

    Article  CAS  PubMed  Google Scholar 

  24. Bhatia M. Hematopoiesis from human embryonic stem cells. Ann N Y Acad Sci 2007;1106:219–222.

    Article  CAS  PubMed  Google Scholar 

  25. Wang L. Endothelial and hematopoietic cell fate of human embryonic stem cells. Trends Cardiovasc Med 2006;16:89–94.

    Article  PubMed  Google Scholar 

  26. Keller GM, Webb S, Kennedy M. Hematopoietic development of ES cells in culture. Methods Mol Med 2002;63:209–230.

    PubMed  Google Scholar 

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Authors and Affiliations

  1. Department of Stem Cell Biology, School of Medicine, Konkuk University, 120-1 Neungdong-ro, Gwangjin-gu, Seoul, 05030, Korea

    Chang-Hyun Gil, Joseph Seo, Jong-Jin Choi, Hana Kim, Sung-Hwan Moon & Hyung-Min Chung

  2. Department of Biomedical Science, CHA University, Seongnam, Korea

    Byeong-Seong Ki & Youngsok Choi

  3. Center for Biomaterials, Korea Institute of Science and Technology, Seoul, Korea

    In-Gul Kim & Kwideok Park

  4. Department of Food Bioscience, Research Institute for Biomedical & Health Science, College of Biomedical & Health Science, Konkuk University, Chungju, Korea

    A.-Ra Jung & Won-Young Lee

Authors
  1. Chang-Hyun Gil
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  2. Byeong-Seong Ki
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  3. Joseph Seo
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  4. Jong-Jin Choi
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  5. Hana Kim
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  6. In-Gul Kim
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  7. A.-Ra Jung
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  8. Won-Young Lee
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  9. Youngsok Choi
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  10. Kwideok Park
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  11. Sung-Hwan Moon
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  12. Hyung-Min Chung
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Corresponding authors

Correspondence to Sung-Hwan Moon or Hyung-Min Chung.

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These authors contributed equally to this work.

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Gil, CH., Ki, BS., Seo, J. et al. Directing human embryonic stem cells towards functional endothelial cells easily and without purification. Tissue Eng Regen Med 13, 274–283 (2016). https://doi.org/10.1007/s13770-016-9076-3

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  • DOI: https://doi.org/10.1007/s13770-016-9076-3

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