The role of non-linear viscoelastic hydrogel mechanics in cell culture and transduction

Abstract

The mechanical complexity of the extracellular matrix (ECM) is central to how cells sense and respond to their environment, yet hydrogel design has often focused narrowly on stiffness. Emerging evidence highlights the importance of viscoelastic stress relaxation and plasticity in cell mechanotransduction. However, a key aspect remains underexplored: non-linear viscoelasticity, where stress relaxation and plasticity depend on the magnitude of applied stress or strain. In this perspective, we examine how such non-linear mechanical behaviors manifest in widely used hydrogels and discuss their biological relevance. We present experimental approaches, including oscillatory shear rheology, to detect non-linear viscoelastic effects, and introduce mathematical modeling approaches to interpret these behaviors. We find evidence in literature that several hydrogels commonly used in cell culture exhibit non-linear viscoelasticity occurring at stress and strain levels relevant to cell-generated forces. Specifically, both softening and stiffening hydrogels were found to exhibit accelerated stress relaxation or increased plasticity due to nonlinear viscoelasticity. By viewing non-linearity as a tunable design parameter, future hydrogel systems may better recapitulate the dynamic mechanical feedback loops cells experience in native tissues. This perspective encourages a paradigm shift in biomaterial design, integrating non-linear viscoelasticity into the next generation of ECM-mimetic hydrogels for cell culture and regenerative applications.

Keywords: Cell biology; Hydrogels; Mathematical modelling; Mechanotransduction; Non-linear viscoelasticity; Rheology.

Graphical abstract

Fig. 1

A) rheometer control and measured parameters for strain sweeps to determine stress-strain response, step-strains to determine viscoelastic stress relaxation, and creep-recovery experiments to determine viscoplasticity of materials. B) Schematic overview of linear and non-linear elasticity as well as linear viscoelasticity showing the same stress relaxation at two different strains or the same strain at two different points in time and non-linear viscoelasticity showing strain-dependent stress relaxation.

Fig. 2

A) Typical stress-strain response of strain-softening hydrogels indicating critical strain γ_c. The illustrations of hydrogels are “Created in Biorender. Sacco, P (2025) https://BioRender.com/w5tefc5 and https://BioRender.com/ncb153l ”. B). Strain sweep of strain-softening hydrogels depicting G′ and G″ as a function of strain indicating the linear viscoelastic regime (LVER) and γ_c. The yellow regime indicates the strain range typically induced by cells. C) Normalized stress relaxation over time at increasing strain for strain-softening hydrogels depicted in B). Both replotted from Ref. [45] D) Plasticity of a set of alginate hydrogels designed to exhibit equivalent stress relaxation but altering plasticity, including non-linear viscoelastic hydrogels with increasing plasticity as a function of strain. Reprinted from Ref. [20], Copyright 2020 National Academy of Sciences. E) Typical stress-strain response of strain-stiffening hydrogels indicating the differential modulus K’. The illustration of hydrogels are “Created in Biorender. Sacco, P (2025) https://BioRender.com/tyd1a8j”. F). K′ as a function of stress for a strain-stiffening hydrogel with K’ = G′ in the LVER and power law increase K’ = σ^m beyond critical stress σ_c. Replotted from Ref. [55]. The yellow regime indicates the stress range corresponding to traction stresses typically exerted by cells, while patterned areas correspond to peak stress observed at cell protrusions or during collective cell migration or mitosis. G) Normalized stress relaxation over time at increasing strain for strain-stiffening collagen hydrogels. Reprinted from Ref. [68], Copyright 2016 National Academy of Sciences. H) Plasticity as a function of creep stress and time for various hydrogels commonly used cell culture in cell culture with strain-stiffening collagen hydrogels exhibiting non-linear viscoelasticity. Reprinted from Ref. [16], (Copyright 2016), with permission from Elsevier. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3

Cartoon. In living tissues, cells are embedded in a 3D microenvironment consisting of an intricate hydrogel network of structural proteins such as collagen and proteoglycans that form the extracellular matrix (ECM). Cells interact closely with the ECM and sense its biophysical properties while generating stress/strain cycles through polymerization/depolymerization of cytoskeletal components or volume expansion. At low stress, cells sense the immediate resistance of the ECM, i.e., linear elasticity or stiffness. The cells can produce an almost immediate, but transient, local non-linear stiffening due to the alignment of the ECM fibers. However, when the stress persists the ECM responds in a time-dependent manner (viscoelasticity). The ECM can eventually undergo plastic deformation when weak bonds in the ECM unbind, allowing the cells to remodel, or when ECM proteins molecularly slide, leading to an arrangement of the ECM by cell contraction. This results in a non-linear softening that causes a diffuse change in the ECM network. We propose that repeated cycles of polymerization/depolymerization of cytoskeletal components generating the same stress/strain on different time scales or gradually increasing their magnitude may lead to non-linear viscoelasticity effects, with enhanced stress relaxation and increased plasticity of the ECM. Created in BioRender. Sacco, P. (2025) https://BioRender.com/apwytrm.