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Generation of kidney organoids from human pluripotent stem cells

Abstract

The human kidney develops from four progenitor populations—nephron progenitors, ureteric epithelial progenitors, renal interstitial progenitors and endothelial progenitors—resulting in the formation of maximally 2 million nephrons. Until recently, the reported methods differentiated human pluripotent stem cells (hPSCs) into either nephron progenitor or ureteric epithelial progenitor cells, consequently forming only nephrons or collecting ducts, respectively. Here we detail a protocol that simultaneously induces all four progenitors to generate kidney organoids within which segmented nephrons are connected to collecting ducts and surrounded by renal interstitial cells and an endothelial network. As evidence of functional maturity, proximal tubules within organoids display megalin-mediated and cubilin-mediated endocytosis, and they respond to a nephrotoxicant to undergo apoptosis. This protocol consists of 7 d of monolayer culture for intermediate mesoderm induction, followed by 18 d of 3D culture to facilitate self-organizing renogenic events leading to organoid formation. Personnel experienced in culturing hPSCs are required to conduct this protocol.

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Figure 1: Schematic diagram of the time line for generating kidney organoids from hPSCs.
Figure 2: Bright-field images of hPSC differentiation to kidney organoids.
Figure 3: Immunological characterization of structures within kidney organoids.

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Acknowledgements

We are grateful to F. Froemling for experimental support and to E.J. Wolvetang (The University of Queensland, St Lucia, Australia) for providing the CRL1502 clone C32 iPSC line. This research was supported by the National Health and Medical Research Council (NHMRC) of Australia (APP1041277), the Australian Research Council (Stem Cells Australia, SRI110001002) and Organovo. M.H.L. is an NHMRC senior principal research fellow. We also acknowledge the use of the Australian Cancer Research Foundation's (ACRF's) Cancer Biology Imaging Facility at The University of Queensland.

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M.T. and M.H.L. wrote the manuscript. M.T., P.X.E. and H.S.C. performed the experiments.

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Correspondence to Minoru Takasato or Melissa H Little.

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Competing interests

M.T. and M.H.L. are named inventors on a patent relating to this methodology.

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Takasato, M., Er, P., Chiu, H. et al. Generation of kidney organoids from human pluripotent stem cells. Nat Protoc 11, 1681–1692 (2016). https://doi.org/10.1038/nprot.2016.098

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