Evaluating the Effect of Stress Relaxation on Cell-based Cartilage Regeneration in 3D using Hydrogels
Creators: name: Johannsen, Julia
name: Yang, Fan
name: Jones, Sarah
Publisher: name: Stanford Digital Repository
Year: 2024
Subjects: subject: Cartilage
subject: Viscoelasticity
subject: Hydrogels
subject: Biomaterials
subject: Chondrocytes
subject: Metabolism

description: Articular cartilage plays an essential role in the shock-absorbing function of joints and in alleviating forces on underlying bone. Yet, while injury to this tissue is common, it has a poor capacity for self-regeneration due to its low cellularity and avascular nature. Cartilage defects, when left untreated, can significantly increase the risk for osteoarthritis (OA), a disease also characterized by a dysregulation of chondrocyte metabolism. Hydrogels, with their highly tunable properties, are a promising strategy for enhancing cell-based therapies to regenerate cartilage. Cartilage tissue is viscoelastic and demonstrates unique stress relaxation behavior upon mechanical loading. Using alginate hydrogels as a model system, it has been recently reported that increasing stress relaxation in 3D can boost cartilage deposition by juvenile chondrocytes. However, the mechanisms by which tuning stress relaxation modulate chondrocyte metabolism and impact human mesenchymal stem cell (hMSC) chondrogenesis remains largely unknown.  
To fill in the above gap of knowledge, the overall goal of this thesis is to harness ionically-crosslinked alginate hydrogels with tunable stress relaxation to elucidate the role of viscoelasticity in modulating cell-based cartilage regeneration and cell metabolism. Rheological tests confirmed successful fabrication of alginate hydrogels with tunable stress relaxation and constant stiffness. Unexpectedly, no conclusive trend in chondrocyte-based cartilage deposition was discernable between fast and slow stress relaxing hydrogels, a finding inconsistent with previous literature. To better understand and contextualize our results, I further assessed the effect of varying cell medium composition, cell density, and material stiffness on chondrocyte matrix formation and metabolism in alginate hydrogels with tunable stress relaxation. Thus, this research aims to utilize an established alginate hydrogel system with tunable viscoelasticity and to investigate the role of stress relaxation on cartilage regeneration and cell behaviors like chondrocyte metabolism and hMSC chondrogenesis. Towards this goal, the deposition of extracellular matrix components in gels of varied viscoelasticity was analyzed as a first pass at harnessing the material system. Glucose consumption by chondrocytes in hydrogels of varying viscoelasticity was also characterized, shedding light on the role of stress relaxation in modulating cellular metabolism.
Unexpectedly, stress relaxation did not promote greater deposition of extracellular matrix by chondrocytes in gels of 5 kPa in stiffness as previously reported. Instead, chondrocytes in slower stress relaxing hydrogels were found to deposit as much, and in some cases more, cartilage matrix than cells in faster stress relaxing hydrogels. We thus wanted to understand whether chondrocyte-based cartilage deposition in response to faster stress relaxation may be stiffness-or cell media-dependent. Interestingly, when stiffer hydrogels were used, a stronger trend indicating greater matrix deposition in faster stress relaxing groups was observed. In this work, we see that the response of cells to stress relaxation may be dependent on other conditions such as cell media and bulk material stiffness. The results of this thesis help to elucidate the role of viscoelasticity in combination with other biochemical and material cues in driving cell-based matrix formation in 3D hydrogels for stem cell-based tissue engineering, guiding the development of optimal materials for tissue regeneration therapies.