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Spatiotemporal Analyses of Cellular Tractions Describe Subcellular Effect of Substrate Stiffness and Coating

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Abstract

Cells interplay with their environment through mechanical and chemical interactions. To characterize this interplay, endothelial cells were cultured on polyacrylamide hydrogels of varying stiffness, coated with either fibronectin or collagen. We developed a novel analysis technique, complementary to traction force microscopy, to characterize the spatiotemporal evolution of cellular tractions: We identified subpopulations of tractions, termed traction foci, and tracked their magnitude and lifetime. Each focus consists of tractions associated with a local single peak of maximal traction. Individual foci were spread over a larger area in cells cultured on collagen relative to those on fibronectin and exerted higher tractions on stiffer hydrogels. We found that the trends with which forces increased with increasing hydrogel stiffness were different for foci and whole-cell measurements. These differences were explained by the number of foci and their average strength. While on fibronectin multiple short-lived weak foci contributed up to 30% to the total traction on hydrogels with intermediate stiffness, short-lived foci in such a number were not observed on collagen despite the higher tractions. Our approach allows for the use of existing traction force microscopy data to gain insight at the subcellular scale without molecular probes or spatial constraining of cellular tractions.

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Acknowledgments

AIA, DAV, and HVO designed research; AIA, DAV, RS, and MV performed research; DAV and AJP developed analytic tools; AIA and DAV analyzed data; AIA, DAV, AJP, and HVO wrote the manuscript. Ben De Coninck was involved in carrying out additional experiments with the vinculin reporter.

Funding

Funding for this work comes from the European Research Council (FP7/2007-2013)/ ERC Grant Agreement (No. 308223), FWO-Vlaanderen (Grants Nos. G.0821.13, G087018N, and G.0B96.15), KU Leuven internal funding (IDO/13/016), and FWO and EU’s Horizon 2020 research and innovation programme (Marie Skłodowska-Curie Grant Agreement No. 665501).

Conflict of interest

All authors declare that there are no known conflicts of interest associated with this submission.

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Author notes

  1. Alicia Izquierdo-Álvarez and Diego A. Vargas have contributed equally to this work.

Authors and Affiliations

  1. Biomechanics Section, KU Leuven, Celestijnenlaan 300C, 3001, Leuven, Belgium

    Alicia Izquierdo-Álvarez, Diego A. Vargas, Álvaro Jorge-Peñas, Marie-Mo Vaeyens & Hans Van Oosterwyck

  2. MeBioS, KU Leuven, Kasteelpark Arenberg 30, 3001, Leuven, Belgium

    Ramesh Subramani

  3. Prometheus, Division of Skeletal Tissue Engineering, KU Leuven, Herestraat 49, 3000, Leuven, Belgium

    Hans Van Oosterwyck

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  1. Alicia Izquierdo-Álvarez
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Correspondence to Hans Van Oosterwyck.

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Associate Editor Debra T. Auguste oversaw the review of this article.

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Izquierdo-Álvarez, A., Vargas, D.A., Jorge-Peñas, Á. et al. Spatiotemporal Analyses of Cellular Tractions Describe Subcellular Effect of Substrate Stiffness and Coating. Ann Biomed Eng 47, 624–637 (2019). https://doi.org/10.1007/s10439-018-02164-2

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