Abstract
Local mechanical stiffness influences cell behavior, and thus cell culture scaffolds should approximate the stiffness of the tissue type from which the cells are derived. In synthetic hydrogels, this has been difficult to achieve for very soft tissues such as neural. This work presents a method for reducing the stiffness of mechanically and biochemically tunable synthetic poly(ethylene glycol) diacrylate hydrogels to within the soft tissue stiffness regime by altering the organization of the crosslinking sites. A soluble allyl-presenting monomer, which has a higher propensity for chain termination than acrylate monomers, was introduced into the PEG-diacrylate hydrogel precursor solution before crosslinking, resulting in acrylate-allyl competition and a reduction in gel compressive modulus from 5.1 ± 0.48 kPa to 0.32 ± 0.09 kPa. Both allyl monomer concentration and chemical structure were shown to influence the effectiveness of competition and change in stiffness. Fibroblast cells demonstrated a 37% reduction in average cell spread area on the softest hydrogels produced as compared to cells on control hydrogels, while the average percentage of neural cells extending neurites increased by 41% on these hydrogels, demonstrating the potential for this technology to serve as a soft tissue culture system.
Highlights
Cell behavior and phenotype are largely influenced by interactions with their local environment.It has been well established that cells receive signals from biochemical factors in their environment such as extracellular proteins and adhesive ligands
We have previously demonstrated a novel method for decreasing PEGDA hydrogel compressive stiffness to within the neural regime by incorporating a short side chain known as allyloxycarbonyl, or alloc, for short, onto a site on the polymer backbone
We constructed PEGDA-based gels with stiffness from 5.1 ± 0.48 kPa to 0.32 ± 0.09 kPa by implementing a competitive monomer that interferes with acrylate crosslinking to reduce the stiffness of the overall hydrogel network structure without preventing gel formation
Summary
Cell behavior and phenotype are largely influenced by interactions with their local environment.It has been well established that cells receive signals from biochemical factors in their environment such as extracellular proteins and adhesive ligands. The stiffness of the local environment determines the magnitude of the tension created, which subsequently influences cell cytoskeletal arrangement and signal pathway activation, and downstream cell activity [5,6,7]. Adhesion, spreading, and migration are all influenced by matrix stiffness [8,9]. The environmental stiffness of tissue culture systems for both investigative and therapeutic applications can greatly impact the activity and fate of cultured cells. It has been thoroughly demonstrated in the literature that matrix stiffness is influential in systems supporting culture of cells from especially soft tissues such as neural cells. Neural cells demonstrate increased neurite extension in softer environments [10,11,12], and neural stem and progenitor cells favor neurogenesis over astrogenesis
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