Quantifying compressive forces between living cell layers and within tissues using elastic round microgels

Abstract

Increasing evidence shows that mechanical stresses are critical in regulating cell functions, fate, and diseases. However, no methods exist that can quantify isotropic compressive stresses. Here we describe fluorescent nanoparticle-labeled, monodisperse elastic microspheres made of Arg-Gly-Asp-conjugated alginate hydrogels (elastic round microgels, ERMGs). We generate 3D displacements and calculate strains and tractions exerted on an ERMG. Average compressive tractions on an ERMG are 570 Pa within cell layers and 360 Pa in tumor-repopulating cell (TRC) colonies grown in 400-Pa matrices. 3D compressive tractions on a 1.4-kPa ERMG are applied by surrounding cells via endogenous actomyosin forces but not via mature focal adhesions. Compressive stresses are substantially heterogeneous on ERMGs within a uniform cell colony and do not increase with TRC colony sizes. Early-stage zebrafish embryos generate spatial and temporal differences in local normal and shear stresses. This ERMG method could be useful for quantifying stresses in vitro and in vivo.

3D microenvironment stiffness regulates tumor spheroid growth and mechanics via p21 and ROCK

“…[35] Another study used alginate beads loaded with fluorescent nanoparticles to measure compressive stresses within melanoma cell spheroids. [36] To our knowledge, polymeric stress sensors have not been used before to measure growth-induced stress in the environment of a tumor spheroid. The novel approach taken in our study therefore provides valuable insights into how tumor spheroids balance their growth and matrix degradation in response to environment stiffness.…”

Superresolved microparticle traction force microscopy reveals subcellular force patterns in immune cell-target interactions

“…Recent reports have described deformable hydrogel particles that can be used as passive force sensors in multicellular aggregates and tissues 15,16 . Compared to the single microfluidic channel methods chosen previously, our batch particle synthesis method is simpler, and requires little specialized expertise or facilities.…”

“…Critical to achieving the required high spatial resolution and sensitivity is our novel strategy for measuring deformation and calculating forces. Unlike classical TFM, which was recently extended to deformable microparticles (~25 μm) 15 and is based on measuring displacement of tracer particles embedded in hydrogels, our approach is based on measurement of the particle boundary with superresolution (~ 50 nm) accuracy. Although computationally more challenging, the resolution in this strategy is not limited by the tracer-to-tracer spacing (> 1 μm) and the approach is very sensitive (~ 10 Pa) because deformation is measured directly at the point of force application.…”

“…Force sensing using spherical sensors is a new area of exploration and first became possible within tissue, and in vivo, by the introduction of oil droplet-based technologies 13,14 . Recently, deformable hydrogel microparticles (MPs) were also used as force reporters 15 . The use of microgels for MP based traction force microscopy (MP-TFM) shows promise for broad applicability because of the mechanical tunability over orders of magnitude and the potential to measure both shear and normal stresses.…”

“…However, current technologies have lacked the resolution to identify individual subcellular force transmitting structures. Moreover, synthesis of suitable hydrogel MPs for such applications can be rather complex, often requiring specialized expertise in microfluidics 15,16 . Particle properties, especially their size and mechanical properties, must also be optimized for accurate traction measurements 16 .…”