Differentiation of human iPSCs into functional podocytes

Caroline Rauch, Elisabeth Feifel, Georg Kern, Cormac Murphy, Florian Meier, Walther Parson, Mario Beilmann, Paul Jennings, Gerhard Gstraunthaler, Anja Wilmes

Research output: Contribution to JournalArticleAcademicpeer-review

Abstract

Podocytes play a critical role in glomerular barrier function, both in health and disease. However, in vivo terminally differentiated podocytes are difficult to be maintained in in vitro culture. Induced pluripotent stem cells (iPSCs) offer the unique possibility for directed differentiation into mature podocytes. The current differentiation protocol to generate iPSC-derived podocyte-like cells provides a robust and reproducible method to obtain podocyte-like cells after 10 days that can be employed in in vitro research and biomedical engineering. Previous published protocols were improved by testing varying differentiation media, growth factors, seeding densities, and time course conditions. Modifications were made to optimize and simplify the one-step differentiation procedure. In contrast to earlier protocols, adherent cells for differentiation were used, the use of fetal bovine serum (FBS) was reduced to a minimum, and thus ß-mercaptoethanol could be omitted. The plating densities of iPSC stocks as well as the seeding densities for differentiation cultures turned out to be a crucial parameter for differentiation results. Conditionally immortalized human podocytes served as reference controls. iPSC-derived podocyte-like cells showed a typical podocyte-specific morphology and distinct expression of podocyte markers synaptopodin, podocin, nephrin and WT-1 after 10 days of differentiation as assessed by immunofluorescence staining or Western blot analysis. qPCR results showed a downregulation of pluripotency markers Oct4 and Sox-2 and a 9-fold upregulation of the podocyte marker synaptopodin during the time course of differentiation. Cultured podocytes exhibited endocytotic uptake of albumin. In toxicological assays, matured podocytes clearly responded to doxorubicin (Adriamycin™) with morphological alterations and a reduction in cell viability after 48 h of incubation.

Original languageEnglish
Article numbere0203869
Pages (from-to)1-18
Number of pages18
JournalPLoS ONE
Volume13
Issue number9
DOIs
Publication statusPublished - 17 Sep 2018

Fingerprint

Induced Pluripotent Stem Cells
Podocytes
Stem cells
doxorubicin
Doxorubicin
sowing
Biomedical engineering
Mercaptoethanol
fetal bovine serum
cells
Cell culture
Plating
in vitro culture
cell differentiation
growth factors
fluorescent antibody technique
cell viability
albumins
Albumins
Assays

Keywords

  • podocytes
  • iPSCs
  • Differentiation

Cite this

Rauch, C., Feifel, E., Kern, G., Murphy, C., Meier, F., Parson, W., ... Wilmes, A. (2018). Differentiation of human iPSCs into functional podocytes. PLoS ONE, 13(9), 1-18. [e0203869]. https://doi.org/10.1371/journal.pone.0203869
Rauch, Caroline ; Feifel, Elisabeth ; Kern, Georg ; Murphy, Cormac ; Meier, Florian ; Parson, Walther ; Beilmann, Mario ; Jennings, Paul ; Gstraunthaler, Gerhard ; Wilmes, Anja. / Differentiation of human iPSCs into functional podocytes. In: PLoS ONE. 2018 ; Vol. 13, No. 9. pp. 1-18.
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Rauch, C, Feifel, E, Kern, G, Murphy, C, Meier, F, Parson, W, Beilmann, M, Jennings, P, Gstraunthaler, G & Wilmes, A 2018, 'Differentiation of human iPSCs into functional podocytes' PLoS ONE, vol. 13, no. 9, e0203869, pp. 1-18. https://doi.org/10.1371/journal.pone.0203869

Differentiation of human iPSCs into functional podocytes. / Rauch, Caroline; Feifel, Elisabeth; Kern, Georg; Murphy, Cormac; Meier, Florian; Parson, Walther; Beilmann, Mario; Jennings, Paul; Gstraunthaler, Gerhard; Wilmes, Anja.

In: PLoS ONE, Vol. 13, No. 9, e0203869, 17.09.2018, p. 1-18.

Research output: Contribution to JournalArticleAcademicpeer-review

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Rauch C, Feifel E, Kern G, Murphy C, Meier F, Parson W et al. Differentiation of human iPSCs into functional podocytes. PLoS ONE. 2018 Sep 17;13(9):1-18. e0203869. https://doi.org/10.1371/journal.pone.0203869