Modeling Cartilage Extracellular Matrix Stiffness: A Potent Regulator of TGFβ-inducible Chondrocyte Differentiation
The extracellular matrix (ECM) functions hierarchically: macroscopically, it supports the tissue under physiologic loading and microscopically, it acts as a physical cue for the native cell population. Combining these perspectives of an orthopedic tissue, like cartilage, provides comprehensive knowledge of the tissue as a whole. In this dissertation, PA gels were adapted to study cartilage ECM to understand its function at the tissue level and as a physical cue in the cellular microenvironment. Layered PA gels were used to emulate the stratified material properties of articular cartilage and develop a mathematical model that describes the loading behavior of intact stratified materials. This model, when applied to articular cartilage, led to the finding that osteoarthritis results in a loss of stratified architecture and an increase in the homogeneity of cartilage ECM. Homogeneous PA gels were used in cellular mechanosensing studies as a physical cue to determine the effect of ECM stiffness on chondrocyte differentiation. Chondrocyte gene expression in ATDC5 cells, murine chondrocytes, and mesenchymal stem cells were specifically induced on substrates that mimicked the stiffness of articular cartilage ECM. Addition of exogenous TGFβ to cartilage-like substrates induced a synergistic induction in chondrocyte specific gene expression. Chondroinduction on cartilage-like substrates requires autocrine TGFβ1 expression. Smad3 phosphorylation, nuclear localization, and transcriptional activity are also induced on cartilage-like substrates. When TGFβ is added exogenously, synergistic induction of chondrocyte gene expression becomes Smad3 independent, acting instead through the p38 MAPK pathway. Combining macroscopic and microscopic perspectives of cartilage ECM from this dissertation may be parlayed into novel therapies and tissue engineering strategies.