Probing Single-Cell Macrophage Polarization and Heterogeneity Using Thermo-Reversible Hydrogels in Droplet-Based Microfluidics

B. M. Tiemeijer; B. M. Tiemeijer; M. W. D. Sweep; M. W. D. Sweep; J. J. F. Sleeboom; J. J. F. Sleeboom; J. J. F. Sleeboom; K. J. Steps; K. J. Steps; J. F. van Sprang; J. F. van Sprang; J. F. van Sprang; P. De Almeida; R. Hammink; R. Hammink; P. H. J. Kouwer; A. I. P. M. Smits; A. I. P. M. Smits; J. Tel; J. Tel

doi:10.3389/fbioe.2021.715408

Frontiers in Bioengineering and Biotechnology (Oct 2021)

Probing Single-Cell Macrophage Polarization and Heterogeneity Using Thermo-Reversible Hydrogels in Droplet-Based Microfluidics

B. M. Tiemeijer,
B. M. Tiemeijer,
M. W. D. Sweep,
M. W. D. Sweep,
J. J. F. Sleeboom,
J. J. F. Sleeboom,
J. J. F. Sleeboom,
K. J. Steps,
K. J. Steps,
J. F. van Sprang,
J. F. van Sprang,
J. F. van Sprang,
P. De Almeida,
R. Hammink,
R. Hammink,
P. H. J. Kouwer,
A. I. P. M. Smits,
A. I. P. M. Smits,
J. Tel,
J. Tel

Affiliations

B. M. Tiemeijer: Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
B. M. Tiemeijer: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
M. W. D. Sweep: Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
M. W. D. Sweep: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
J. J. F. Sleeboom: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
J. J. F. Sleeboom: Microsystems, Department of Mechanical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
J. J. F. Sleeboom: Laboratory of Soft Tissue Engineering and Biomechanics, Department Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
K. J. Steps: Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
K. J. Steps: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
J. F. van Sprang: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
J. F. van Sprang: Laboratory of Soft Tissue Engineering and Biomechanics, Department Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
J. F. van Sprang: Laboratory of Chemical Biology, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
P. De Almeida: Department of System Chemistry, Institute for Molecules and Materials, Radboud University, Nijmegen, Netherlands
R. Hammink: Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, Netherlands
R. Hammink: Oncode Institute, Radboud University Medical Center, Nijmegen, Netherlands
P. H. J. Kouwer: Department of System Chemistry, Institute for Molecules and Materials, Radboud University, Nijmegen, Netherlands
A. I. P. M. Smits: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands
A. I. P. M. Smits: Laboratory of Soft Tissue Engineering and Biomechanics, Department Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
J. Tel: Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands
J. Tel: Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands

DOI: https://doi.org/10.3389/fbioe.2021.715408
Journal volume & issue: Vol. 9

Abstract

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Human immune cells intrinsically exist as heterogenous populations. To understand cellular heterogeneity, both cell culture and analysis should be executed with single-cell resolution to eliminate juxtacrine and paracrine interactions, as these can lead to a homogenized cell response, obscuring unique cellular behavior. Droplet microfluidics has emerged as a potent tool to culture and stimulate single cells at high throughput. However, when studying adherent cells at single-cell level, it is imperative to provide a substrate for the cells to adhere to, as suspension culture conditions can negatively affect biological function and behavior. Therefore, we combined a droplet-based microfluidic platform with a thermo-reversible polyisocyanide (PIC) hydrogel, which allowed for robust droplet formation at low temperatures, whilst ensuring catalyzer-free droplet gelation and easy cell recovery after culture for downstream analysis. With this approach, we probed the heterogeneity of highly adherent human macrophages under both pro-inflammatory M1 and anti-inflammatory M2 polarization conditions. We showed that co-encapsulation of multiple cells enhanced cell polarization compared to single cells, indicating that cellular communication is a potent driver of macrophage polarization. Additionally, we highlight that culturing single macrophages in PIC hydrogel droplets displayed higher cell viability and enhanced M2 polarization compared to single macrophages cultured in suspension. Remarkably, combining phenotypical and functional analysis on single cultured macrophages revealed a subset of cells in a persistent M1 state, which were undetectable in conventional bulk cultures. Taken together, combining droplet-based microfluidics with hydrogels is a versatile and powerful tool to study the biological function of adherent cell types at single-cell resolution with high throughput.

Published in Frontiers in Bioengineering and Biotechnology

ISSN: 2296-4185 (Online)
Publisher: Frontiers Media S.A.
Country of publisher: Switzerland
LCC subjects: Technology: Chemical technology: Biotechnology
Website: http://www.frontiersin.org/bioengineering_and_biotechnology

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