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Liver Regeneration Using Cultured Liver Bud

Part of the Methods in Molecular Biology book series (MIMB,volume 1597)

Abstract

Here, we describe a protocol to develop a three-dimensional (3D) liver bud-like tissue from human iPSCs in vitro. This method mainly consists of two parts: (1) hepatic endoderm (HE) differentiation from human iPSCs in 2D culture and (2) co-culturing iPSC-HE with endothelial and mesenchymal cells. First, iPSCs were differentiated into definitive endoderm (DE) cells, and the DE cells were differentiated into HE cells, which were then co-cultured with endothelial cells and mesenchymal cells on Matrigel-coated plastic plates or micropattern plates. The cells rapidly condensed to generate 3D tissue masses. We named these iPSC liver buds (iPSC-LBs) because they resemble the developing liver bud from the perspective of gene expression, cell proliferation, and cell proportion. This liver bud culture system provides a novel approach for future clinical applications, for drug development, and as a tool for studying human development.

Key words

  • Human-induced pluripotent stem cells
  • Liver bud
  • Three-dimensional culture
  • Self-organization
  • Multicellular interaction
  • Regenerative medicine
  • Drug development

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References

  1. Mclaren A (2001) Ethical and social considerations of stem cell research. Nature 414:129–131. doi:10.1038/35102194

    CAS  CrossRef  PubMed  Google Scholar 

  2. Takahashi K, Tanabe K, Ohnuki M et al (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861–872. doi:10.1016/j.cell.2007.11.019

    CAS  CrossRef  PubMed  Google Scholar 

  3. Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676. doi:10.1016/j.cell.2006.07.024

    CAS  CrossRef  PubMed  Google Scholar 

  4. Miura K, Okada Y, Aoi T et al (2009) Variation in the safety of induced pluripotent stem cell lines. Nat Biotechnol 27:743–745. doi:10.1038/nbt.1554. pii: nbt.1554 [pii]\r

    CAS  CrossRef  PubMed  Google Scholar 

  5. Si-Tayeb K, Noto FK, Nagaoka M et al (2010) Highly efficient generation of human hepatocyte-like cells from induced pluripotent stem cells. Hepatology 51:297–305. doi:10.1002/hep.23354

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  6. Si-Tayeb K, Lemaigre FP, Duncan SA (2010) Organogenesis and development of the liver. Dev Cell 18:175–189. doi:10.1016/j.devcel.2010.01.011

    CAS  CrossRef  PubMed  Google Scholar 

  7. Zaret KS, Grompe M (2008) Generation and regeneration of cells of the liver and pancreas. Science 322:1490–1494. doi:10.1126/science.1161431

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  8. Franklin V, Khoo PL, Bildsoe H et al (2008) Regionalisation of the endoderm progenitors and morphogenesis of the gut portals of the mouse embryo. Mech Dev 125:587–600. doi:10.1016/j.mod.2008.04.001

    CAS  CrossRef  PubMed  Google Scholar 

  9. Arterbery AS, Bogue CW (2014) Endodermal and mesenchymal cross talk: a crossroad for the maturation of foregut organs. Pediatr Res 75:120–126. doi:10.1038/pr.2013.201

    CrossRef  PubMed  Google Scholar 

  10. Lokmane L, Haumaitre C, Garcia-Villalba P et al (2008) Crucial role of vHNF1 in vertebrate hepatic specification. Development 135:2777–2786. doi:10.1242/dev.023010

    CAS  CrossRef  PubMed  Google Scholar 

  11. Sekine K, Chen Y-R, Kojima N et al (2007) Foxo1 links insulin signaling to C/EBPalpha and regulates gluconeogenesis during liver development. EMBO J 26:3607–3615

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  12. Kajiwara M, Aoi T, Okita K et al (2012) Donor-dependent variations in hepatic differentiation from human-induced pluripotent stem cells. Proc Natl Acad Sci USA 109:12538–12543. doi:10.1073/pnas.1209979109

    CAS  CrossRef  PubMed  PubMed Central  Google Scholar 

  13. Grapin-Botton A (2008) Endoderm specification. Stem Book 3–4. doi: 10.3824/stembook.1.30.1

  14. Matsuno K, Mae SI, Okada C, et al (2016) Redefining definitive endoderm subtypes by robust induction of human induced pluripotent stem cells. Differentiation 1–10. doi: 10.1016/j.diff.2016.04.002

  15. Sekine K, Takebe T, Suzuki Y et al (2012) Highly efficient generation of definitive endoderm lineage from human induced pluripotent stem cells. Transplant Proc 44:1127–1129. doi:10.1016/j.transproceed.2012.03.001

    CAS  CrossRef  PubMed  Google Scholar 

  16. Collin de l’Hortet A, Takeishi K, Guzman-Lepe J et al (2016) Liver-regenerative transplantation: regrow and reset. Am J Transplant 16:1688–1696. doi:10.1111/ajt.13678

    CrossRef  PubMed  Google Scholar 

  17. Takebe T, Sekine K, Enomura M et al (2013) Vascularized and functional human liver from an iPSC-derived organ bud transplant. Nature 499:481–484. doi:10.1038/nature12271

    CAS  CrossRef  PubMed  Google Scholar 

  18. Takebe T, Zhang R-R, Koike H et al (2014) Generation of a vascularized and functional human liver from an iPSC-derived organ bud transplant. Nat Protoc 9:396–409. doi:10.1038/nprot.2014.020

    CAS  CrossRef  PubMed  Google Scholar 

  19. Takebe T, Sekine K, Suzuki Y et al (2012) Self-organization of human hepatic organoid by recapitulating organogenesis in vitro. Transplant Proc 44:1018–1020. doi:10.1016/j.transproceed.2012.02.007

    CAS  CrossRef  PubMed  Google Scholar 

  20. Takebe T, Koike N, Sekine K et al (2012) Generation of functional human vascular network. Transplant Proc 44:1130–1133. doi:10.1016/j.transproceed.2012.03.039

    CAS  CrossRef  PubMed  Google Scholar 

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Correspondence to Hideki Taniguchi .

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Sekine, K., Takebe, T., Taniguchi, H. (2017). Liver Regeneration Using Cultured Liver Bud. In: Tsuji, T. (eds) Organ Regeneration. Methods in Molecular Biology, vol 1597. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6949-4_15

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  • DOI: https://doi.org/10.1007/978-1-4939-6949-4_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6947-0

  • Online ISBN: 978-1-4939-6949-4

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