The study of intercellular signalling networks requires organotypic microscale systems that facilitate the culture, conditioning and manipulation of cells. Here, we describe a reconfigurable microfluidic cell-culture system that facilitates the assembly of three-dimensional tissue models by stacking layers that contain preconditioned microenvironments. By using principles of open and suspended microfluidics, the Stacks system is easily assembled or disassembled to provide spatial and temporal manoeuvrability in two-dimensional and three-dimensional assays of multiple cell types, enabling the modelling of sequential paracrine-signalling events, such as tumour-cell-mediated differentiation of macrophages and macrophage-facilitated angiogenesis. We used Stacks to recapitulate the in vivo observation that different prostate cancer tissues polarize macrophages with distinct gene-expression profiles of pro-inflammatory and anti-inflammatory cytokines. Stacks also enabled us to show that these two types of macrophages signal distinctly to endothelial cells, leading to blood vessels with different morphologies. Our proof-of-concept experiments exemplify how Stacks can efficiently model multicellular interactions and highlight the importance of spatiotemporal specificity in intercellular signalling.
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This work is funded by a National Science Foundation grant (EFRI-MKIS), University of Wisconsin Carbone Cancer Center Cancer Center Support Grant (P30 CA014520), Research Training in Hematology T32 (NIH T32 HL07899), NIH R01EB010039 BRG, NIH R01 CA185251, NIH K12 DK100022, DOD Prostate Cancer Research Program (W81XWH-16-0543), the Arnold and Mabel Beckman Foundation (Beckman Young Investigator Award), and the University of Washington. We thank D. Kosoff for helpful discussions and assistance with PCR, and undergraduate student B. Horman for facilitating experiments.
The authors have potential conflicts of interest related to technologies presented here: J.Y. (Stacks to the Future), E.B. (Tasso, Salus Discovery and Stacks to the Future), T.E.d.G. (Stacks to the Future and Lynx Biosciences), A.B.T. (Stacks to the Future), and D.J.B. (Bellbrook Labs, Tasso, Stacks to the Future, Lynx Biosciences, Onexio Biosystems and Salus Discovery). However, none of these companies supported this work.
Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Figs. 1–6, methods and captions for Supplementary Videos 1–3.
Stacks layers are open microfluidic devices that can be operated with a pipette.
Stacks devices can be assembled to enable diffusion.
Stacks devices are reconfigurable and enable diffusion across layers.
ImageJ macros for image processing.
SolidWorks files for the Stack devices.
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Yu, J., Berthier, E., Craig, A. et al. Reconfigurable open microfluidics for studying the spatiotemporal dynamics of paracrine signalling. Nat Biomed Eng 3, 830–841 (2019). https://doi.org/10.1038/s41551-019-0421-4
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