Quantifying force transmission through fibroblasts: changes of traction forces under external shearing

Steven Huth,Johannes W Blumberg,Jan Lammerding,Dimitri Probst,Ulrich S Schwarz,Christine Selhuber-Unkel

doi:10.1007/s00249-021-01576-8

Abstract

Mammalian cells have evolved complex mechanical connections to their microenvironment, including focal adhesion clusters that physically connect the cytoskeleton and the extracellular matrix. This mechanical link is also part of the cellular machinery to transduce, sense and respond to external forces. Although methods to measure cell attachment and cellular traction forces are well established, these are not capable of quantifying force transmission through the cell body to adhesion sites. We here present a novel approach to quantify intracellular force transmission by combining microneedle shearing at the apical cell surface with traction force microscopy at the basal cell surface. The change of traction forces exerted by fibroblasts to underlying polyacrylamide substrates as a response to a known shear force exerted with a calibrated microneedle reveals that cells redistribute forces dynamically under external shearing and during sequential rupture of their adhesion sites. Our quantitative results demonstrate a transition from dipolar to monopolar traction patterns, an inhomogeneous distribution of the external shear force to the adhesion sites as well as dynamical changes in force loading prior to and after the rupture of single adhesion sites. Our strategy of combining traction force microscopy with external force application opens new perspectives for future studies of force transmission and mechanotransduction in cells.

Highlights

Cells exert forces to interact with their surroundings and have the striking ability to react to externally applied forces and mechanical cues by a process called mechanotransduction (Jaalouk and Lammerding 2009; Petridou et al 2017; Roca-Cusachs et al 2017)
Focal adhesion clusters grow in response to external shearing (Riveline 2001; Paul et al 2008) which might help cells to withstand shear forces, e.g., forces exerted by the blood flow on endothelial cells (Davies 1995; Perrault 2015)
Traction force microscopy (TFM) has become an established tool to quantify forces exerted by single cells or cell layers to the underlying substrate, which has deepened our understanding of cell migration, mechanotransduction and cell–matrix interaction (Lo et al 2000; Schwarz and Soiné 2015; Style 2014; Kronenberg 2017; Sabass et al 2008; 3 Vol.:(0123456789)

Summary

Introduction

Cells exert forces to interact with their surroundings and have the striking ability to react to externally applied forces and mechanical cues by a process called mechanotransduction (Jaalouk and Lammerding 2009; Petridou et al 2017; Roca-Cusachs et al 2017). We present a new tool that combines TFM with externally applied mechanical stimulation by microneedle shearing This setting allows to quantify cellular force transmission by measuring how cells distribute an external welldefined shear force to their adhesion sites. We advanced current TFM procedures to create a novel procedure that analyzes traction forces in the presence of an external force monopole This new force transmission assay is a versatile technique that is complementary to existing methods, as it can be combined with other techniques such as AFM to broaden our understanding of the interplay of cellular biomechanics and adhesion. C shows a plot of the shear force versus the microneedle bending for each frame of the experiment

Results and discussion

Conclusion

Materials and methods