Abstract
An organoid can be defined as a three-dimensional organ-like structure formed from organ-specific progenitor cells. Organ progenitor cells were empirically found to self-organize three-dimensional tissues when they were aggregated and cultivated in vitro. While this nature power of progenitor cells has an amazing potential to recreate artificial organs in vitro, there had been difficulty to apply this technology to human organs due to the inaccessibility to human progenitor cells until human-induced pluripotent stem cell (hiPSC) was invented by Takahashi and Yamanaka in 2007. As embryonic stem cells do, hiPSCs also have pluripotency to give rise to any organs/tissues cell types, including the kidney, via directed differentiation. Here, we provide a detailed protocol for generating kidney organoids using human pluripotent stem cells. The protocol differentiates human pluripotent stem cells into the posterior primitive streak. This is followed by the simultaneous induction of posterior and anterior intermediate mesoderm that are subsequently aggregated and undergo self-organization into the kidney organoid. Such kidney organoids are comprised of all anticipated kidney cell types including nephrons segmented into the glomerulus, proximal tubule, loop of Henle, and distal tubule as well as the collecting duct, endothelial network, and renal interstitium.
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References
Bertram JF, Douglas-Denton RN, Diouf B et al (2011) Human nephron number: implications for health and disease. Pediatr Nephrol 26(9):1529–1533
Sajithlal G, Zou D, Silvius D et al (2005) Eya1 acts as a critical regulator for specifying the metanephric mesenchyme. Dev Biol 284:323–336
Xu J, Wong EYM, Cheng C et al (2014) Eya1 interacts with Six2 and Myc to regulate expansion of the nephron progenitor pool during nephrogenesis. Dev Cell 31:434–447
Vega QC, Worby CA, Lechner MS et al (1996) Glial cell line-derived neurotrophic factor activates the receptor tyrosine kinase RET and promotes kidney morphogenesis. Proc Natl Acad Sci U S A 93:10657–10661
Majumdar A, Vainio S, Kispert A et al (2003) Wnt11 and Ret/Gdnf pathways cooperate in regulating ureteric branching during metanephric kidney development. Development 130:3175–3185
Carroll TJ, Park J, Hayashi S et al (2005) Wnt9b plays a central role in the regulation of mesenchymal to epithelial transitions underlying organogenesis of the mammalian urogenital system. Dev Cell 9:283–292
Barak H, Huh S-H, Chen S et al (2012) FGF9 and FGF20 maintain the stemness of nephron progenitors in mice and man. Dev Cell 22:1191–1207
Tamplin OJ, Cox BJ, Rossant J (2011) Integrated microarray and ChIP analysis identifies multiple Foxa2 dependent target genes in the notochord. Dev Biol 360:415–425
Mugford JW, Sipilä P, McMahon JA et al (2008) Osr1 expression demarcates a multi-potent population of intermediate mesoderm that undergoes progressive restriction to an Osr1-dependent nephron progenitor compartment within the mammalian kidney. Dev Biol 324:88–98
Takasato M, Er PX, Chiu HS et al (2015) Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. Nature 526:564–568
Tam PPL, Loebel DAF (2007) Gene function in mouse embryogenesis: get set for gastrulation. Nat Rev Genet 8:368–381
Takasato M, Er PX, Becroft M et al (2014) Directing human embryonic stem cell differentiation towards a renal lineage generates a self-organizing kidney. Nat Cell Biol 16:118–126
Sumi T, Tsuneyoshi N, Nakatsuji N et al (2008) Defining early lineage specification of human embryonic stem cells by the orchestrated balance of canonical Wnt/−catenin, Activin/Nodal and BMP signaling. Development 135:2969–2979
Davis RP, Ng ES, Costa M et al (2008) Targeting a GFP reporter gene to the MIXL1 locus of human embryonic stem cells identifies human primitive streak-like cells and enables isolation of primitive hematopoietic precursors. Blood 111:1876–1884
Ng ES, Davis R, Stanley EG et al (2008) A protocol describing the use of a recombinant protein-based, animal product-free medium (APEL) for human embryonic stem cell differentiation as spin embryoid bodies. Nat Protoc 3:768–776
Colvin JS, Feldman B, Nadeau JH et al (1999) Genomic organization and embryonic expression of the mouse fibroblast growth factor 9 gene. Dev Dyn 216:72–88
Takasato M, Little MH (2015) The origin of the mammalian kidney: implications for recreating the kidney in vitro. Development 142:1937–1947
Saxen L (1987) Organogenesis of the kidney. Cambridge University Press, Cambridge
Giuliani S, Perin L, Sedrakyan S et al (2008) Ex vivo whole embryonic kidney culture: a novel method for research in development, regeneration and transplantation. J Urol 179:365–370
Gupta IR, Lapointe M, Yu OH (2003) Morphogenesis during mouse embryonic kidney explant culture. Kidney Int 63:365–376
Acknowledgments
M.L. is a National Health and Medical Research Council Senior Principal Research Fellow. This work was supported by the NHMRC and the Australian Research Council. M.C.R.I. is supported by the Victorian Government’s Operational Infrastructure Support Program.
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Takasato, M., Little, M.H. (2017). Making a Kidney Organoid Using the Directed Differentiation of Human Pluripotent Stem Cells. 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_14
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DOI: https://doi.org/10.1007/978-1-4939-6949-4_14
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