Generation of 3D Soluble Signal Gradients in Cell-Laden Hydrogels Using Passive Diffusion.

Autor: Ahrens L; Institute for Macromolecular Chemistry, Hermann Staudinger Haus, University of Freiburg, Stefan-Meier-Str. 31, 79104, Freiburg, Germany.; BIOSS Centre for Signalling Studies, University of Freiburg, Schänzlestraße 18, 79104, Freiburg, Germany., Tanaka S; Computational Biology Group, D-BSSE, ETH Zürich, Mattenstrasse 26, 4058, Basel, Switzerland., Vonwil D; Institute for Macromolecular Chemistry, Hermann Staudinger Haus, University of Freiburg, Stefan-Meier-Str. 31, 79104, Freiburg, Germany., Christensen J; Institute for Macromolecular Chemistry, Hermann Staudinger Haus, University of Freiburg, Stefan-Meier-Str. 31, 79104, Freiburg, Germany.; BIOSS Centre for Signalling Studies, University of Freiburg, Schänzlestraße 18, 79104, Freiburg, Germany., Iber D; Computational Biology Group, D-BSSE, ETH Zürich, Mattenstrasse 26, 4058, Basel, Switzerland., Shastri VP; Institute for Macromolecular Chemistry, Hermann Staudinger Haus, University of Freiburg, Stefan-Meier-Str. 31, 79104, Freiburg, Germany.; BIOSS Centre for Signalling Studies, University of Freiburg, Schänzlestraße 18, 79104, Freiburg, Germany.
Jazyk: angličtina
Zdroj: Advanced biosystems [Adv Biosyst] 2019 Jan; Vol. 3 (1), pp. e1800237. Date of Electronic Publication: 2018 Nov 14.
DOI: 10.1002/adbi.201800237
Abstrakt: Soluble signal gradients play an important role in organ patterning, cell migration, and differentiation. Currently, signal gradients in 2D cell culture are realized using microfluidics and here cells are exposed to high and nonphysiological shear stress. Tissue morphogenesis (organogenesis) however occurs in 3D and therefore there is a need for simple and practical systems to impose gradients to cells dispersed in 3D matrix. Herein, a 3D gradient generator based on passive diffusion elements that recapitulates interstitial flow and is capable of imposing predictable gradients over long length scales (6 mm) lasting up to 48 h to cells dispersed in a hydrogel environment is reported. Using recombinant human WNT3A (rhWNT3A), the spatiotemporal activation of the canonical WNT pathway in human epithelial kidney cells and human mesenchymal stems cells expressing a green fluorescence protein reporter on a transcription factor/lymphoid enhancer-binding factor (TCF/LEF) promoter is demonstrated. By refining computation models based on experimental findings, the diffusion coefficient of rhWNT3A in presence of human cells in 3D is determined. Furthermore, the formation of rhBMP4 gradients is visualized using immunohistochemistry by staining for phospho-SMAD1/5, the downstream targets of the bone morphogenetic protein (BMP) pathway. The simplicity of the gradient generator is expected to spur its adoption in studying developmental biology paradigms in vitro.
(© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)
Databáze: MEDLINE
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