(637d) Mechanical Characterization of Elastic and Viscoelastic Polyacrylamide Hydrogels for Cell-Substrate Interaction Studies

Polyacrylamide hydrogels (PAH) are soft biomaterials often used in bioengineering and tissue engineering to mimic the mechanical properties of native extracellular matrices (ECM) and to study how cells mechanosense and respond to varied environments. This is partly due to the ability to tune the elastic properties of PAH to within the range relevant to soft tissues (Young's modulus of 1 – 40 kPa) by adjusting their chemical compositions. For affine gels, two (of three) elastic constants -- Young's modulus (E), the shear modulus (G), and the Poisson's ratio (ν) -- allow describing the purely elastic response to external forces. Recent work using PAH has demonstrated how one may independently tune the elastic storage (G') and the viscous loss (G") moduli of these model ECMs. Since PAH are porous with high water content, we hypothesize that viscoelastic and poroelastic gel properties may also impact and regulate cell mechanobiology. Thus viscoelastic relaxation times and the frequency-dependent complex moduli of PAH, for time-scales and frequencies relevant to cell-substrate interactions, need to be characterized. This is particularly important in quantitative studies, like traction force microscopy (TFM) where cell induced substrate deformations are directly visualized, quantified, and then used with appropriate constitutive models for the gel to estimate focal adhesion related stress fields. Constitutive models typically employ linear elasticity theory for affine materials and require knowledge of E and ν. The Young's modulus is commonly measured by compressional indentation tests such as micro-indentation, followed by applying a standard model such as the Hertz model to interpret force-indentation profiles. While the need for accurate estimation of Young's modulus is well recognized, the importance of correctly estimating the Poisson’s ratio and the viscoelastic relaxation time constants is often overlooked. For instance, for computational ease, especially in TFM calculations, hydrogels are treated as incompressible (ν = 0.5). In this study, gels with graded properties are made by using varying ratios of acrylamide (monomer) and bis-acrylamide (cross-linker). The mechanical properties of these gels are then characterized, and their interdependence is studied via microindentation, shear rheology, and compressional rheology. We highlight how experimentally determined values for elastic constants depends on the deformation rate. Viscoelastic relaxation time scales (from shear rheology) and the poroelastic relaxation times (from compression experiments) are related to the chemical composition of the gel. The water content of the gel, pore size, and the Poisson’s ratio are also directly inferred from experiments. Taken together, our suite of experiments enables us to completely characterize static and dynamic mechanical properties of PAH relevant to cell-substrate studies. These fully characterized PAH can be used for mechanobiology experiments to quantitatively investigate cell migration, cell-traction forces, and morphological changes with the correct and full complement of mechanical properties.