Designing Spatiotemporally Tunable Viscoelastic Hyaluronic Acid Hydrogels to Study Cell Mechanobiology During Fibrosis

The ECM is highly dynamic and is constantly regulated through cell-cell and cell-matrix interactions. ECM dysregulation can trigger various signaling cascades and promote deviant cell behaviors and disease progression. Fibrosis is a heterogeneous pathological scarring outcome of many diseases that is characterized by progressive matrix stiffening and decreasing viscoelasticity. Treatments have been largely unsuccessful due to a lack of suitable test systems for probing molecular mechanisms involved in fibrogenesis. Additionally, most hydrogel models fail to display tissue-relevant time-dependent properties such as stress relaxation, which has shown to be a critical regulator of cell phenotype. Engineered hydrogels with tunable properties has become a powerful method to study the mechanoregulatory role of mechanical and biochemical cues on cell behaviors. Overall, the goal of this thesis is to develop a class of mechanically dynamic and spatiotemporally heterogeneous hydrogels with precise control over cell-instructive inputs, allowing recapitulation of both normal and diseased microenvironments.

For this work, we used phototunable hyaluronic acid (HA)-based hydrogels that enable independent tuning of stiffness, viscoelasticity, and adhesive ligand presentation to understand how physical and chemical cues collectively influence cell behaviors in fibrosis. Chapter 3 characterizes the time-dependent properties of ex vivo tissue, such as stress relaxation and frequency-dependent behavior, to direct the design of ECM-mimetic biomaterials. Chapter 4 illustrates the development of an in vitro fibrosis model to study how stiffness, viscoelasticity, and heterogeneity influences fibroblast response. Chapter 5 uses engineered fibronectin adhesive fragments to understand the individual and combined roles of stiffness, viscoelasticity, and adhesion on cell spreading, actin stress fiber formation, and focal adhesion maturation. Finally, Chapter 6 explores how viscoelastic cues impact cell behaviors in 3D cultures. Altogether, these studies present a framework for engineering instructive hydrogel platforms that offer greater insight into the role that complex matrix properties play in regulating cell mechanobiology in the context of fibrosis progression.