The generation of kidney organoids by differentiation of human pluripotent cells to ureteric bud progenitor–like cells


This protocol presents recently developed methodologies for the differentiation of human pluripotent stem cells (hPSCs) into ureteric bud (UB) progenitor–like cells. Differentiation of human PSCs to UB progenitor–like cells allows for the generation of chimeric kidney cultures in which the human cells can self-assemble into chimeric 3D structures in combination with embryonic mouse kidney cells over a period of 18 d. UB progenitor–like cells are generated by a two-step process that combines in vitro commitment of human PSCs, whether embryonic stem cells (ESCs) or induced PSCs (iPSCs), under chemically defined culture conditions, with ex vivo cultures for the induction of 3D organogenesis. The models described here provide new opportunities for investigating human kidney development, modeling disease, evaluating regenerative medicine strategies, as well as for toxicology studies.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Schematic representation of the differentiation procedure.
Figure 2: Overview of the reaggregation assay.
Figure 3: E12.5 mouse kidney aggregates formed by different starting cell numbers.
Figure 4: Morphology of mouse and chimeric aggregate cultures.
Figure 5: Embryonic renal marker expression of chimeric aggregates.
Figure 6: Adult renal marker expression in aggregate culture.


  1. 1

    Takebe, T. et al. Generation of a vascularized and functional human liver from an iPSC-derived organ bud transplant. Nat. Protoc. 9, 396–409 (2014).

    CAS  Article  Google Scholar 

  2. 2

    Lancaster, M.A. et al. Cerebral organoids model human brain development and microcephaly. Nature 501, 373–379 (2013).

    CAS  Article  Google Scholar 

  3. 3

    Nakano, T. et al. Self-formation of optic cups and storable stratified neural retina from human ESCs. Cell Stem Cell 10, 771–785 (2012).

    CAS  Article  Google Scholar 

  4. 4

    Hockemeyer, D. et al. Genetic engineering of human pluripotent cells using TALE nucleases. Nat. Biotechnol. 29, 731–734 (2011).

    CAS  Article  Google Scholar 

  5. 5

    Shen, B. et al. Efficient genome modification by CRISPR-Cas9 nickase with minimal off-target effects. Nat. Methods 11, 399–402 (2014).

    CAS  Article  Google Scholar 

  6. 6

    Liu, G.H. et al. Targeted gene correction of laminopathy-associated LMNA mutations in patient-specific iPSCs. Cell Stem Cell 8, 688–694 (2011).

    CAS  Article  Google Scholar 

  7. 7

    Xia, Y. et al. Directed differentiation of human pluripotent cells to ureteric bud kidney progenitor-like cells. Nat. Cell Biol. 15, 1507–1515 (2013).

    CAS  Article  Google Scholar 

  8. 8

    Taguchi, A. et al. Redefining the in vivo origin of metanephric nephron progenitors enables generation of complex kidney structures from pluripotent stem cells. Cell Stem Cell 14, 53–67 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9

    Takasato, M. et al. Directing human embryonic stem cell differentiation towards a renal lineage generates a self-organizing kidney. Nat. Cell Biol. 16, 118–126 (2014).

    CAS  Article  Google Scholar 

  10. 10

    Raya, A. et al. Disease-corrected haematopoietic progenitors from Fanconi anaemia induced pluripotent stem cells. Nature 460, 53–59 (2009).

    CAS  Article  Google Scholar 

  11. 11

    Kabgani, N. et al. Primary cultures of glomerular parietal epithelial cells or podocytes with proven origin. PLoS ONE 7, e34907 (2012).

    CAS  Article  Google Scholar 

  12. 12

    Shankland, S.J., Pippin, J.W., Reiser, J. & Mundel, P. Podocytes in culture: past, present, and future. Kidney Int. 72, 26–36 (2007).

    CAS  Article  Google Scholar 

  13. 13

    Little, M.H. & McMahon, A.P. Mammalian kidney development: principles, progress, and projections. Cold Spring Harb. Perspect. Biol. 4, pii: a008300 (2012).

    Article  Google Scholar 

  14. 14

    Batchelder, C.A., Lee, C.C., Matsell, D.G., Yoder, M.C. & Tarantal, A.F. Renal ontogeny in the rhesus monkey (Macaca mulatta) and directed differentiation of human embryonic stem cells towards kidney precursors. Differentiation 78, 45–56 (2009).

    CAS  Article  Google Scholar 

  15. 15

    Vigneau, C. et al. Mouse embryonic stem cell-derived embryoid bodies generate progenitors that integrate long term into renal proximal tubules in vivo. J. Am. Soc. Nephrol. 18, 1709–1720 (2007).

    CAS  Article  Google Scholar 

  16. 16

    Bruce, S.J. et al. In vitro differentiation of murine embryonic stem cells toward a renal lineage. Differentiation 75, 337–349 (2007).

    CAS  Article  Google Scholar 

  17. 17

    Kim, D. & Dressler, G.R. Nephrogenic factors promote differentiation of mouse embryonic stem cells into renal epithelia. J. Am. Soc. Nephrol. 16, 3527–3534 (2005).

    CAS  Article  Google Scholar 

  18. 18

    Davies, J.A., Unbekandt, M., Ineson, J., Lusis, M. & Little, M.H. Dissociation of embryonic kidney followed by re-aggregation as a method for chimeric analysis. Methods Mol. Biol. 886, 135–146 (2012).

    CAS  Article  Google Scholar 

  19. 19

    Unbekandt, M. & Davies, J.A. Dissociation of embryonic kidneys followed by reaggregation allows the formation of renal tissues. Kidney Int. 77, 407–416 (2010).

    Article  Google Scholar 

  20. 20

    Chang, C.H. & Davies, J.A. An improved method of renal tissue engineering, by combining renal dissociation and reaggregation with a low-volume culture technique, results in development of engineered kidneys complete with loops of Henle. Nephron Exp. Nephrol. 121, e79–85 (2012).

    CAS  Article  Google Scholar 

  21. 21

    Sebinger, D.D. et al. A novel, low-volume method for organ culture of embryonic kidneys that allows development of cortico-medullary anatomical organization. PLoS ONE 5, e10550 (2010).

    Article  Google Scholar 

  22. 22

    Amit, M. & Itskovitz-Eldor, J. Atlas of Human Pluripotent Stem Cells (Humana Press, 2012).

  23. 23

    Costantini, F., Watanabe, T., Lu, B., Chi, X. & Srinivas, S. Dissection of embryonic mouse kidney, culture in vitro, and imaging of the developing organ. Cold Spring Harb. Protoc. 2011 10.1101/pdb.prot5613 (2011).

  24. 24

    Ludwig, T.E. et al. Feeder-independent culture of human embryonic stem cells. Nat. Methods 3, 637–646 (2006).

    CAS  Article  Google Scholar 

Download references


We thank M. Schwarz for administrative support. We thank J. Kasuboski from the Waitt Advanced Biophotonics Core at the Salk Institute for Biological Studies for help with imaging processing. Y.X. was partially supported by the California Institute for Regenerative Medicine (CIRM) through a CIRM Training grant. I.S.-M. was partially supported by a Nomis Foundation postdoctoral fellowship. Work in the laboratory of J.C.I.B. was supported by grants from the G. Harold and Leila Y. Mathers Charitable Foundation and The Leona M. and Harry B. Helmsley Charitable Trust (2012-PG-MED002).

Author information




Y.X., I.S.-M., E.N., C.R.E., J.M.C. and J.C.I.B. designed all experiments and developed the methodologies presented here. Y.X., I.S.-M., E.N. and J.C.I.B. wrote the manuscript.

Corresponding author

Correspondence to Juan Carlos Izpisua Belmonte.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Xia, Y., Sancho-Martinez, I., Nivet, E. et al. The generation of kidney organoids by differentiation of human pluripotent cells to ureteric bud progenitor–like cells. Nat Protoc 9, 2693–2704 (2014).

Download citation

Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing