Journal
NATURE PROTOCOLS
Volume 9, Issue 2, Pages 396-409Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nprot.2014.020
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Funding
- Ministry of Education, Culture, Sports, Science, and Technology of Japan [24106510, 24689052, 22390260, 21249071, 25253079]
- Research Center Network for Realization of Regenerative Medicine
- Strategic Promotion of Innovative Research and Development program (S-innovation) of the JST [62890004]
- Takeda Science Foundation
- Japan IDDM network
- Yokohama Foundation for Advanced Medical Science
- Grants-in-Aid for Scientific Research [24689052, 26106721, 24659593, 22390260, 21249071, 24106510, 24659785, 25253079] Funding Source: KAKEN
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Generation of functional and vascularized organs from human induced pluripotent stem cells (iPSCs) will facilitate our understanding of human developmental biology and disease modeling, hopefully offering a drug-screening platform and providing novel therapies against end-stage organ failure. Here we describe a protocol for the in vitro generation of a 3D liver bud from human iPSC cultures and the monitoring of further hepatic maturation after transplantation at various ectopic sites. iPSC-derived specified hepatic cells are dissociated and suspended with endothelial cells and mesenchymal stem cells. These mixed cells are then plated onto a presolidified matrix, and they form a 3D spherical tissue mass termed a liver bud (iPSC-LB) in 1-2 d. To facilitate additional maturation, 4-d-old iPSC-LBs are transplanted in the immunodeficient mouse. Live imaging has identified functional blood perfusion into the preformed human vascular networks. Functional analyses show the appearance of multiple hepatic functions in a chronological manner in vivo.
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