Introducing Dynamicity: Engineering Stress Relaxation Into Hydrogels Via Thiol-Ene Modified Alginate for Mechanobiological in vitro Modeling of the Cornea.

Developing biomaterials for corneal repair and regeneration is an important area of research, as the cornea is a vital and highly specialized tissue that plays a crucial role in maintaining clear vision. Corneal keratocytes, the primary cell type of the corneal stroma, are sensitive to their surrounding mechanical environment. Altering stiffness of the cellular microenvironment can upregulate actin stress fibre production and focal adhesions, as well as alter the keratocytes' phenotypic markers. Stiffness however, is a static mechanical property that does not truly capture the dynamic environment of in vivo tissue. The human cornea has been characterized as a stiff tissue, with viscoelastic properties that have been measured before but it is not well understood how these properties influence cellular interactions embedded within this tissue. The hypothesis of this paper is that the cornea has time-dependent mechanical properties, much like other tissues of the body those properies can be recapitulated via stress relaxation in potential therapeutic matrices. The stress relaxation properties of the cornea were uncovered by nanoindentation, where the relaxation of the tissue achieved 15% relaxation within 10 s. Using a specially formulated alginate-PEG and alginate-norbornene mixture, the researchers tuned the hydrogel matrix's dynamicity through photoinitiated norbornene-norbornene dimerization spanning a range of 30 s to 10 minutes. Subsequently, the impact of these viscoelastic adjustments on the hydrogel were assessed with human primary corneal keratocytes (HPCK) especially their survival and gene expression. Primary corneal keratocytes showed a decrease in αSMA expression on slower relaxing hydrogels and did not stain for these contractile proteins in any of the hydrogels. Keratocytes cultured on slower relaxing hydrogels, closer in relaxation profile to that of the decellularized cornea, displayed more filopodia than those on fast relaxing hydrogels, similarly to their in vivo phenotype, suggesting the slower relaxing material influenced cells to adopt a more native-like phenotype. This in vitro model can be used to determine optimal stress relaxation for multiple cell types, including primary corneal keratocytes to control contractile tissue formation. By combining optimization of stress relaxation alongside the commonly assessed property of stiffness, this platform not only provides a more accurate tool for in vitro study of cell behaviour, but may reduce mechanical mismatch between native tissues and implanted constructs. This article is protected by copyright. All rights reserved.