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Basic Research

Manipulation of Nephron-Patterning Signals Enables Selective Induction of Podocytes from Human Pluripotent Stem Cells

Yoshimura, Yasuhiro; Taguchi, Atsuhiro; Tanigawa, Shunsuke; Yatsuda, Junji; Kamba, Tomomi; Takahashi, Satoru; Kurihara, Hidetake; Mukoyama, Masashi; Nishinakamura, Ryuichi

1Department of Kidney Development, Institute of Molecular Embryology and Genetics, and

Departments of 2Nephrology and

4Urology, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan;

3Department of Genome Regulation, Max Planck Institute for Molecular Genetics, Berlin, Germany;

5Department of Anatomy and Embryology, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan; and

6Department of Anatomy and Life Structure, Juntendo University School of Medicine, Tokyo, Japan

Correspondence: Dr. Ryuichi Nishinakamura, Department of Kidney Development, Institute of Molecular Embryology and Genetics, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan, or Dr. Atsuhiro Taguchi, Department of Genome Regulation, Max Planck Institute for Molecular Genetics, Ihnestraße 63-73, 14195 Berlin, Germany. E-mail: [email protected] or [email protected]

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Abstract

Significance Statement

Recent progress in stem cell biology has enabled researchers to induce nephron progenitor cells (NPCs) and kidney organoids from pluripotent stem cells (PSCs). However, shepherding NPC differentiation toward a specific nephron segment remains a challenge. The authors observed the effects of various growth factors in cultured mouse embryonic NPCs during three phases of the differentiation process, demonstrating that phase-specific manipulation of Wnt and Tgf-β signaling is critical for podocyte differentiation. Using this insight into the nephron-patterning process, they were able to selectively induce human PSC-derived podocytes with molecular, morphologic, and functional characteristics of human podocytes. This novel protocol will facilitate accessibility to human podocytes, and these PSC-derived podocytes are expected to serve as a valuable resource in kidney research.

Background

Previous research has elucidated the signals required to induce nephron progenitor cells (NPCs) from pluripotent stem cells (PSCs), enabling the generation of kidney organoids. However, selectively controlling differentiation of NPCs to podocytes has been a challenge.

Methods

We investigated the effects of various growth factors in cultured mouse embryonic NPCs during three distinct steps of nephron patterning: from NPC to pretubular aggregate, from the latter to epithelial renal vesicle (RV), and from RV to podocyte. We then applied the findings to human PSC-derived NPCs to establish a method for selective induction of human podocytes.

Results

Mouse NPC differentiation experiments revealed that phase-specific manipulation of Wnt and Tgf-β signaling is critical for podocyte differentiation. First, optimal timing and intensity of Wnt signaling were essential for mesenchymal-to-epithelial transition and podocyte differentiation. Then, inhibition of Tgf-β signaling supported domination of the RV proximal domain. Inhibition of Tgf-β signaling in the third phase enriched the podocyte fraction by suppressing development of other nephron lineages. The resultant protocol enabled successful induction of human podocytes from PSCs with >90% purity. The induced podocytes exhibited global gene expression signatures comparable to those of adult human podocytes, had podocyte morphologic features (including foot process–like and slit diaphragm–like structures), and showed functional responsiveness to drug-induced injury.

Conclusions

Elucidation of signals that induce podocytes during the nephron-patterning process enabled us to establish a highly efficient method for selective induction of human podocytes from PSCs. These PSC-derived podocytes show molecular, morphologic, and functional characteristics of podocytes, and offer a new resource for disease modeling and nephrotoxicity testing.

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Erratum

Yoshimura Y, Taguchi A, Tanigawa S, Yatsuda J, Kamba T, Takahasi S, Kurihara H, Mukoyama M, Nishinakamura R: Manipulation of Nephron-Patterning Signals Enables Selective Induction of Podocytes from Human Pluripotent Stem Cells. J Am Soc Nephrol 30: 304–321, 2019; DOI: https://doi.org/10.1681/ASN.2018070747

In the Methods section of this article, there are two locations where the concentrations are listed incorrectly in µg/ml when they should be in ng/ml. On pages 305 and 307, “in serum-free differentiation medium containing Fgf9 (10 µg/ml; R&D Systems)” should be “in serum-free differentiation medium containing Fgf9 (10 ng/ml; R&D Systems).” We apologize for the errors and for the inconvenience they may have caused.

See original article, “Manipulation of Nephron-Patterning Signals Enables Selective Induction of Podocytes from Human Pluripotent Stem Cells”, in Vol. 30, Iss. 2, on pages .

Journal of the American Society of Nephrology. 33(1):253, January 2022.