Covalently cross-linked hydrogels: Mechanisms of nonlinear viscoelasticity

Gelatin-based hydrogels have been widely used in tissue engineering, three-dimensional cell culture, drug delivery, and cell therapy. The mechanical behaviour of hydrogels combined with their chemical properties determines their functionality and efficacy. With respect to the mechanical behaviour of hydrogels, the vast majority of publications have reported their linear viscoelastic response. However, for practical conditions in the body, these materials experience large deformations beyond the linear viscoelastic limit. Herein, to mimic practical conditions and to evaluate the mechanical response of the hydrogels subjected to large deformations, we report inter- and intra-cycle nonlinear viscoelastic behaviour of a gelatin methacryloyl (GelMA) hydrogel with different concentrations of the hydrogel precursor (10%-20% [w/v]) under large amplitude oscillatory shear deformation. To achieve this, we used a novel technique by chemically bonding the hydrogels to treated glass slides, which were attached to the oscillating metal plates using a double-sided tape to alleviate any error arising from wall slip during rheological measurements. The results show that the elasticity of the covalently cross-linked hydrogels at large deformations obeys a nonlinear force-extension law and that the viscous intra-cycle nonlinearity at moderate deformations stems from the dual cross-linked (DC; i.e., physical and chemical) nature of the GelMA hydrogel. It was also shown that viscoelastic parameters can be tuned by the concentration of the hydrogel precursor, that is, yield stress increased from 2.6-7.1 kPa, critical strain amplitude decreased from gamma(0) = 100%-70%, and the onset of inter-cycle nonlinearity shifted from gamma(0) = 50%-20% upon increasing the concentration of the hydrogel precursor. These insights have important implications for the rational development of hydrogel-based biomaterials to design biocompatible scaffolds in tissue engineering applications.