Derivation of Functionally Mature Eosinophils from Human Pluripotent Stem Cells

  • Ya Zhou
  • Xu Pan
  • Wenyu Yang
  • Yanzheng Gu
  • Bin Mao
  • Mowen Lai
  • Wencui Sun
  • Shu Huang
  • Tatsutoshi Nakahata
  • Feng MaEmail author
Part of the SpringerBriefs in Stem Cells book series (BRIEFSSTEM, volume 6)


The in vitro development of functionally mature blood cells from human pluripotent stem cells [hPSCs; including human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs)] has proven ideal way to gain information during human early embryonic/fetal hematopoiesis, which never can be mimicked in other species. We recently established an efficient method to produce large quantity of pure and functionally mature eosinophils from hPSCs. The method includes majorly three steps: (1) induction of hematopoietic stem/progenitor cells by coculturing hPSCs with mAGM-3 or mFL stromal cells; (2) large expansion of hPSC-derived hematopoietic stem/progenitor cells and inducing differentiation toward myeloid lineage; and (3) directed differentiation of myeloid progenitor cells into functionally mature eosinophils. The eosinophils induced from hPSCs in our system showed similar morphology and surface marker expression with those in peripheral blood through May-Grϋnwald-Giemsa staining, transmission electron microscopy (TEM) analysis, flow cytometric analysis (FACS), RT-PCR analysis, and immunofluorescent staining. Furthermore, chemotactic migration and degranulation ability have confirmed the maturity and function of these hPSC-derived eosinophils. The induction of eosinophils from hPSCs provides us with a perfect model to study the germination, development, differentiation, and maturation of human eosinophiles, which has not been well defined yet. It also provides novel approach to develop patient-tailored therapies by iPSCs for severe allergic diseases as well as deficiencies in early innate immunity. In this review, we will describe the details of methodology for generating these functionally mature eosinophils from hPSCs and the related assay for their function and maturation.


Human pluripotent stem cells (hPSCs) Eosinophils Differentiation Hematopoiesis mAGM-3 


  1. 1.
    Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM. Embryonic stem cell lines derived from human blastocysts. Science. 1998;282:1145–7.CrossRefPubMedGoogle Scholar
  2. 2.
    Ma F, Ebihara Y, Umeda K, Sakai H, Hanada S, Zhang H, Zaike Y, Tsuchida E, Nakahata T, Nakauchi H. Generation of functional erythrocytes from human embryonic stem cell-derived definitive hematopoiesis. Proc Natl Acad Sci. 2008;105:13087–92.PubMedCentralCrossRefPubMedGoogle Scholar
  3. 3.
    Lu S-J, Feng Q, Park JS, Vida L, Lee B-S, Strausbauch M, Wettstein PJ, Honig GR, Lanza R. Biologic properties and enucleation of red blood cells from human embryonic stem cells. Blood. 2008;112:4475–84.PubMedCentralCrossRefPubMedGoogle Scholar
  4. 4.
    Saeki K, Saeki K, Nakahara M, Matsuyama S, Nakamura N, Yogiashi Y, Yoneda A, Koyanagi M, Kondo Y, Yuo A. A feeder-free and efficient production of functional neutrophils from human embryonic stem cells. Stem Cells. 2009;27:59–67.CrossRefPubMedGoogle Scholar
  5. 5.
    Yokoyama Y, Suzuki T, Sakata-Yanagimoto M, Kumano K, Higashi K, Takato T, Kurokawa M, Ogawa S, Chiba S. Derivation of functional mature neutrophils from human embryonic stem cells. Blood. 2009;113:6584–92.CrossRefPubMedGoogle Scholar
  6. 6.
    Gaur M, Kamata T, Wang S, Moran B, Shattil SJ, Leavitt AD. Megakaryocytes derived from human embryonic stem cells: a genetically tractable system to study Megakaryocytopoiesis and integrin function. J Thromb Haemost. 2005;4:436–42.CrossRefGoogle Scholar
  7. 7.
    Takayama N, Nishikii H, Usui J, Tsukui H, Sawaguchi A, Hiroyama T, Eto K, Nakauchi H. Generation of functional platelets from human embryonic stem cells in vitro via ES-sacs, VEGF-promoted structures that concentrate hematopoietic progenitors. Blood. 2008;111:5298–306.CrossRefPubMedGoogle Scholar
  8. 8.
    Choi K-D, Vodyanik MA, Slukvin II. Generation of mature human myelomonocytic cells through expansion and differentiation of pluripotent stem cell–derived lin–CD34+ CD43+ CD45+ progenitors. J Clin Investig. 2009;119:2818–29.PubMedCentralCrossRefPubMedGoogle Scholar
  9. 9.
    Slukvin II, Vodyanik MA, Thomson JA, Gumenyuk ME, Choi K-D. Directed differentiation of human embryonic stem cells into functional dendritic cells through the myeloid pathway. J Immunol. 2006;176:2924–32.CrossRefPubMedGoogle Scholar
  10. 10.
    Woll PS, Martin CH, Miller JS, Kaufman DS. Human embryonic stem cell-derived NK cells acquire functional receptors and cytolytic activity. J Immunol. 2005;175:5095–103.CrossRefPubMedGoogle Scholar
  11. 11.
    Kovarova M, Latour AM, Chason KD, Tilley SL, Koller BH. Human embryonic stem cells: a source of mast cells for the study of allergic and inflammatory diseases. Blood. 2010;115:3695–703.PubMedCentralCrossRefPubMedGoogle Scholar
  12. 12.
    Vodyanik MA, Bork JA, Thomson JA, Slukvin II. Human embryonic stem cell–derived CD34+ cells: efficient production in the coculture with OP9 stromal cells and analysis of lymphohematopoietic potential. Blood. 2005;105:617–26.CrossRefPubMedGoogle Scholar
  13. 13.
    Timmermans F, Velghe I, Vanwalleghem L, De Smedt M, Van Coppernolle S, Taghon T, Moore HD, Leclercq G, Langerak AW, Kerre T. Generation of T cells from human embryonic stem cell-derived hematopoietic zones. J Immunol. 2009;182:6879–88.CrossRefPubMedGoogle Scholar
  14. 14.
    Giarratana M-C, Kobari L, Lapillonne H, Chalmers D, Kiger L, Cynober T, Marden MC, Wajcman H, Douay L. Ex vivo generation of fully mature human red blood cells from hematopoietic stem cells. Nat Biotechnol. 2004;23:69–74.CrossRefPubMedGoogle Scholar
  15. 15.
    Olsen AL, Stachura DL, Weiss MJ. Designer blood: creating hematopoietic lineages from embryonic stem cells. Blood. 2006;107:1265–75.PubMedCentralCrossRefPubMedGoogle Scholar
  16. 16.
    Ng ES, Davis RP, Azzola L, Stanley EG, Elefanty AG. Forced aggregation of defined numbers of human embryonic stem cells into embryoid bodies fosters robust, reproducible hematopoietic differentiation. Blood. 2005;106:1601–3.CrossRefPubMedGoogle Scholar
  17. 17.
    Wang ZZ, Au P, Chen T, Shao Y, Daheron LM, Bai H, Arzigian M, Fukumura D, Jain RK, Scadden DT. Endothelial cells derived from human embryonic stem cells form durable blood vessels in vivo. Nat Biotechnol. 2007;25:317–8.CrossRefPubMedGoogle Scholar
  18. 18.
    Kaufman DS, Hanson ET, Lewis RL, Auerbach R, Thomson JA. Hematopoietic colony-forming cells derived from human embryonic stem cells. Proc Natl Acad Sci. 2001;98:10716–21.PubMedCentralCrossRefPubMedGoogle Scholar
  19. 19.
    Qiu C, Hanson E, Olivier E, Inada M, Kaufman DS, Gupta S, Bouhassira EE. Differentiation of human embryonic stem cells into hematopoietic cells by coculture with human fetal liver cells recapitulates the globin switch that occurs early in development. Exp Hematol. 2005;33:1450–8.CrossRefPubMedGoogle Scholar
  20. 20.
    Ledran MH, Krassowska A, Armstrong L, Dimmick I, Renström J, Lang R, Yung S, Santibanez-Coref M, Dzierzak E, Stojkovic M. Efficient hematopoietic differentiation of human embryonic stem cells on stromal cells derived from hematopoietic niches. Cell Stem Cell. 2008;3:85–98.CrossRefPubMedGoogle Scholar
  21. 21.
    Yu J, Vodyanik MA, He P, Slukvin II, Thomson JA. Human embryonic stem cells reprogram myeloid precursors following cell–cell fusion. Stem Cells. 2006;24:168–76.CrossRefPubMedGoogle Scholar
  22. 22.
    Matsuoka S, Tsuji K, Hisakawa H, Xu M-J, Ebihara Y, Ishii T, Sugiyama D, Manabe A, Tanaka R, Ikeda Y. Generation of definitive hematopoietic stem cells from murine early yolk sac and paraaortic splanchnopleures by aorta-gonad-mesonephros region–derived stromal cells. Blood. 2001;98:6–12.CrossRefPubMedGoogle Scholar
  23. 23.
    Ma F, Kambe N, Wang D, Shinoda G, Fujino H, Umeda K, Fujisawa A, Ma L, Suemori H, Nakatsuji N. Direct development of functionally mature tryptase/chymase double-positive connective tissue-type mast cells from primate embryonic stem cells. Stem Cells. 2008;26:706–14.CrossRefPubMedGoogle Scholar
  24. 24.
    Lee H, Shamy GA, Elkabetz Y, Schofield CM, Harrsion NL, Panagiotakos G, Socci ND, Tabar V, Studer L. Directed differentiation and transplantation of human embryonic stem cell-derived motoneurons. Stem Cells. 2007;25:1931–9.CrossRefPubMedGoogle Scholar
  25. 25.
    Tian X, Woll PS, Morris JK, Linehan JL, Kaufman DS. Hematopoietic engraftment of human embryonic stem cell-derived cells is regulated by recipient innate immunity. Stem Cells. 2006;24:1370–80.CrossRefPubMedGoogle Scholar
  26. 26.
    Ma F, Wang D, Hanada S, Ebihara Y, Kawasaki H, Zaike Y, Heike T, Nakahata T, Tsuji K. Novel method for efficient production of multipotential hematopoietic progenitors from human embryonic stem cells. Int J Hematol. 2007;85:371–9.CrossRefPubMedGoogle Scholar
  27. 27.
    Ma F, Gu Y, Nishihama N, Yang W, Yasuhiro E, Tsuji K. Differentiation of human embryonic and induced pluripotent stem cells into blood cells in coculture with murine stromal cells. In: Ye K, Jin S, editors. Human embryonic and induced pluripotent stem cells (pp. 321–335). Totowa: Humana Press (2012). doi: 10.1007/978-1-61779-267-0_23. ISBN 978-1-61779-266-3 (Published by Springer Science + Business Modia, LLC 2011).
  28. 28.
    Xu MJ, Tsuji K, Ueda T, Mukouyama YS, Hara T, Yang FC, Ebihara Y, Matsuoka S, Manabe A, Kikuchi A, Ito M, Miyajima A, Nakahata T. Stimulation of mouse and human primitive hematopoiesis by murine embryonic aorta-gonad-mesonephros-derived stromal cell lines. Blood. 1998;92:2032–40.PubMedGoogle Scholar
  29. 29.
    Shamri R, Xenakis JJ, Spencer LA. Eosinophils in innate immunity: an evolving story. Cell Tissue Res. 2011;343:57–83.PubMedCentralCrossRefPubMedGoogle Scholar
  30. 30.
    Lacy P, Rosenberg HF, Walsh GM. Eosinophil overview: structure, biological properties, and key functions. In Eosinophils. Berlin: Springer; 2014. pp 1–12.Google Scholar
  31. 31.
    Rosenberg HF, Dyer KD, Foster PS. Eosinophils: changing perspectives in health and disease. Nat Rev Immunol. 2012;13:9–22.PubMedCentralCrossRefPubMedGoogle Scholar
  32. 32.
    Fulkerson PC, Rothenberg ME. Targeting eosinophils in allergy, inflammation and beyond. Nat Rev Drug Discovery. 2013;12:117–29.CrossRefPubMedGoogle Scholar
  33. 33.
    Crescenzi B, Chase A, La Starza R, Beacci D, Rosti V, Galli A, Specchia G, Martelli M, Vandenberghe P, Cools J. FIP1L1-PDGFRA in chronic eosinophilic leukemia and BCR-ABL1 in chronic myeloid leukemia affect different leukemic cells. Leukemia. 2007;21:397–402.CrossRefPubMedGoogle Scholar
  34. 34.
    Bochner BS, Gleich GJ. What targeting eosinophils has taught us about their role in diseases. J Allergy Clin Immunol. 2010;126:16–25.PubMedCentralCrossRefPubMedGoogle Scholar
  35. 35.
    Hruz P, Straumann A, Bussmann C, Heer P, Simon H-U, Zwahlen M, Beglinger C, Schoepfer AM. Escalating incidence of eosinophilic esophagitis: a 20-year prospective, population-based study in Olten County, Switzerland. J Allergy Clin Immunol. 2011;128(1349–1350):e1345.Google Scholar
  36. 36.
    Bohm M, Malik Z, Sebastiano C, Thomas R, Gaughan J, Kelsen S, Richter JE. Mucosal eosinophilia: prevalence and racial/ethnic differences in symptoms and endoscopic findings in adults over 10 years in an urban hospital. J Clin Gastroenterol. 2012;46:567–74.CrossRefPubMedGoogle Scholar
  37. 37.
    Boyce JA, Friend D, Matsumoto R, Austen KF, Owen WF. Differentiation in vitro of hybrid eosinophil/basophil granulocytes: autocrine function of an eosinophil developmental intermediate. J Exp Med. 1995;182:49–57.CrossRefPubMedGoogle Scholar

Copyright information

© The Author(s) 2015

Authors and Affiliations

  • Ya Zhou
    • 1
  • Xu Pan
    • 1
  • Wenyu Yang
    • 2
  • Yanzheng Gu
    • 3
  • Bin Mao
    • 1
  • Mowen Lai
    • 1
  • Wencui Sun
    • 1
  • Shu Huang
    • 1
  • Tatsutoshi Nakahata
    • 4
  • Feng Ma
    • 1
    • 2
    • 3
    • 5
    Email author
  1. 1.Institute of Blood TransfusionChinese Academy of Medical Sciences and Peking Union Medical CollegeChengduChina
  2. 2.State Key Lab of Experimental HematologyChinese Academy of Medical Sciences and Peking Union Medical CollegeTianjinChina
  3. 3.Stem Cell Key Laboratory of Jiangsu ProvinceSuzhou UniversitySuzhouChina
  4. 4.Center for iPS Cell Research and Application (CiRA)Kyoto UniversityKyotoJapan
  5. 5.Research Center for Stem Cell Therapy, Institute of Blood TransfusionChinese Academy of Medical Sciences and Peking Union Medical College (CAMS&PUMC)ChengduChina

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