Sammanfattning

Kidney mesenchyme (KM) and nephron progenitors (NPs) depend on WNT activity, and their culture in vitro requires extensive repertoire of recombinant proteins and chemicals. Here we established a robust, simple culture of mouse KM using a combination of 3D Matrigel and growth media supplemented with Fibroblast Growth Factor 2 (FGF2) and Src inhibitor PP2. This allows dissociated KM to spontaneously self-organize into spheres. To reassess the requirement of WNT activity in KM self-organization and NPs maintenance, cells were cultured with short pulse of high-dose GSK3 beta inhibitor BIO, on a constant low-dose or without BIO. Robust proliferation at 48 hours and differentiation at 1 week were observed in cultures with high BIO pulse. Importantly, dissociated KM cultured without BIO, similarly to that exposed to constant low dose of BIO, maintained NPs up to one week and spontaneously differentiated into nephron tubules at 3 weeks of culture. Our results show that KM is maintained and induced to differentiate in a simple culture system. They also imply that GSK3 beta/WNT-independent pathways contribute to the maintenance and induction of mouse KM. The robust and easy 3D culture enables further characterization of NPs, and may facilitate disease modeling when applied to human cells.

Originalspråkengelska
Artikelnummer13433
TidskriftScientific Reports
Volym 9
Antal sidor10
ISSN2045-2322
DOI
StatusPublicerad - 17 sep 2019
MoE-publikationstypA1 Tidskriftsartikel-refererad

Vetenskapsgrenar

  • 1184 Genetik, utvecklingsbiologi, fysiologi
  • 1182 Biokemi, cell- och molekylärbiologi

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title = "Simple 3D culture of dissociated kidney mesenchyme mimics nephron progenitor niche and facilitates nephrogenesis Wnt-independently",
abstract = "Kidney mesenchyme (KM) and nephron progenitors (NPs) depend on WNT activity, and their culture in vitro requires extensive repertoire of recombinant proteins and chemicals. Here we established a robust, simple culture of mouse KM using a combination of 3D Matrigel and growth media supplemented with Fibroblast Growth Factor 2 (FGF2) and Src inhibitor PP2. This allows dissociated KM to spontaneously self-organize into spheres. To reassess the requirement of WNT activity in KM self-organization and NPs maintenance, cells were cultured with short pulse of high-dose GSK3 beta inhibitor BIO, on a constant low-dose or without BIO. Robust proliferation at 48 hours and differentiation at 1 week were observed in cultures with high BIO pulse. Importantly, dissociated KM cultured without BIO, similarly to that exposed to constant low dose of BIO, maintained NPs up to one week and spontaneously differentiated into nephron tubules at 3 weeks of culture. Our results show that KM is maintained and induced to differentiate in a simple culture system. They also imply that GSK3 beta/WNT-independent pathways contribute to the maintenance and induction of mouse KM. The robust and easy 3D culture enables further characterization of NPs, and may facilitate disease modeling when applied to human cells.",
keywords = "1184 Genetics, developmental biology, physiology, Kidney development, Imaging analysis, 1182 Biochemistry, cell and molecular biology, IN-VITRO PROPAGATION, BRANCHING MORPHOGENESIS, URETERAL BUD, GENERATION, ORGANOIDS, ORGANOGENESIS, INACTIVATION, POPULATION, EXPRESSION, TISSUES",
author = "Arvydas Dapkunas and Ville Rantanen and Yujuan Gui and Maciej Lalowski and Kirsi Sainio and Satu Kuure and Hannu Sariola",
year = "2019",
month = "9",
day = "17",
doi = "10.1038/s41598-019-49526-x",
language = "English",
volume = "9",
journal = "Scientific Reports",
issn = "2045-2322",
publisher = "Nature Publishing Group",

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TY - JOUR

T1 - Simple 3D culture of dissociated kidney mesenchyme mimics nephron progenitor niche and facilitates nephrogenesis Wnt-independently

AU - Dapkunas, Arvydas

AU - Rantanen, Ville

AU - Gui, Yujuan

AU - Lalowski, Maciej

AU - Sainio, Kirsi

AU - Kuure, Satu

AU - Sariola, Hannu

PY - 2019/9/17

Y1 - 2019/9/17

N2 - Kidney mesenchyme (KM) and nephron progenitors (NPs) depend on WNT activity, and their culture in vitro requires extensive repertoire of recombinant proteins and chemicals. Here we established a robust, simple culture of mouse KM using a combination of 3D Matrigel and growth media supplemented with Fibroblast Growth Factor 2 (FGF2) and Src inhibitor PP2. This allows dissociated KM to spontaneously self-organize into spheres. To reassess the requirement of WNT activity in KM self-organization and NPs maintenance, cells were cultured with short pulse of high-dose GSK3 beta inhibitor BIO, on a constant low-dose or without BIO. Robust proliferation at 48 hours and differentiation at 1 week were observed in cultures with high BIO pulse. Importantly, dissociated KM cultured without BIO, similarly to that exposed to constant low dose of BIO, maintained NPs up to one week and spontaneously differentiated into nephron tubules at 3 weeks of culture. Our results show that KM is maintained and induced to differentiate in a simple culture system. They also imply that GSK3 beta/WNT-independent pathways contribute to the maintenance and induction of mouse KM. The robust and easy 3D culture enables further characterization of NPs, and may facilitate disease modeling when applied to human cells.

AB - Kidney mesenchyme (KM) and nephron progenitors (NPs) depend on WNT activity, and their culture in vitro requires extensive repertoire of recombinant proteins and chemicals. Here we established a robust, simple culture of mouse KM using a combination of 3D Matrigel and growth media supplemented with Fibroblast Growth Factor 2 (FGF2) and Src inhibitor PP2. This allows dissociated KM to spontaneously self-organize into spheres. To reassess the requirement of WNT activity in KM self-organization and NPs maintenance, cells were cultured with short pulse of high-dose GSK3 beta inhibitor BIO, on a constant low-dose or without BIO. Robust proliferation at 48 hours and differentiation at 1 week were observed in cultures with high BIO pulse. Importantly, dissociated KM cultured without BIO, similarly to that exposed to constant low dose of BIO, maintained NPs up to one week and spontaneously differentiated into nephron tubules at 3 weeks of culture. Our results show that KM is maintained and induced to differentiate in a simple culture system. They also imply that GSK3 beta/WNT-independent pathways contribute to the maintenance and induction of mouse KM. The robust and easy 3D culture enables further characterization of NPs, and may facilitate disease modeling when applied to human cells.

KW - 1184 Genetics, developmental biology, physiology

KW - Kidney development

KW - Imaging analysis

KW - 1182 Biochemistry, cell and molecular biology

KW - IN-VITRO PROPAGATION

KW - BRANCHING MORPHOGENESIS

KW - URETERAL BUD

KW - GENERATION

KW - ORGANOIDS

KW - ORGANOGENESIS

KW - INACTIVATION

KW - POPULATION

KW - EXPRESSION

KW - TISSUES

U2 - 10.1038/s41598-019-49526-x

DO - 10.1038/s41598-019-49526-x

M3 - Article

VL - 9

JO - Scientific Reports

JF - Scientific Reports

SN - 2045-2322

M1 - 13433

ER -