Fig. 3: LSCZeb1⁺ cells can self-renew and generate functional prostate in vivo.

a Immunostaining of E-cadherin and Vimentin to validate the purity of UGM cells separated from UGS. (Scale bars = 20 μm.). b Prostate tissue generated from 1000 LSCZeb1⁺ prostate cells at 2 months after renal capsule implantation. (implanted cell number: 1000, scale bars = 2 mm.). c Quantification of prostate tissue generation incidence from 1000-cell transplants. d H&E staining of 1000-cell implants shows well differentiated prostate epithelial tubules. (Scale bars = 20  μm.). e Immunostaining of p63, CK5, CK8, Syp, Pbsn, AR, mouse-specific β1-integrin and C57BL/6 donor-specific H-2k^b on sections of 1000-cell implants. (All scale bars = 20 μm.). f, g Immunostaining of frozen sections from LSCZeb1⁻ cell transplants displaying unstructured tissue containing basal cells, rare luminal cells and mouse-β1-integrin⁻ vimentin⁺ rat mesenchymal cells. (Scale bars = 50 μm). h Phase contrast and fluorescent images of the single viable Lineage⁻ Sca-1⁺ CD49f^hi Zeb1⁺ or Lineage⁻ Sca-1⁺ CD49f^hi Zeb1⁻ prostate cell used for single cell renal capsule transplantation experiments in vivo. (Scale bars = 20 μm.). i, Prostate tissue generated from single Lineage⁻ Sca-1⁺ CD49f^hi Zeb1⁺ prostate cell at 2 months after renal capsule implantation. (scale bars = 2 mm). j Quantification of prostate tissue generation incidence from single cell transplants. k H&E staining of single cell implants showing differentiated prostate epithelial tubules. (Scale bars = 20 μm.). l, Staining of RFP, p63, CK5, CK8, Synaptosphysin (Syp), Pbsn, Androgen receptor (AR), β1-integrin and H-2k^b on sections of single cell implants. (The white arrow points to a Syp⁺ cell. Scale bars = 20 μm.). m, n Zeb1⁺ EpCAM⁺ cells harvested from first generation of renal capsule implantation were able to form prostates in secondary and tertiary transplantations. (50 prostate cells per graft). o, p Immunostaining and quantification of frozen sections from serial transplants showing a stable percentage of Zeb1⁺ basal cells during the serial implantation. (n = 3 grafts. Source data are provided as a Source Data file.).