Valvular interstitial cells (VICs), the main cell population within the aortic heart valve, transition from a quiescent fibroblast phenotype to an active phenotype, such as a pro-fibrotic myofibroblast or a pro-calcific osteoblast-like phenotype in response to microenvironmental cues. Maladaptive effects of prolonged VIC activation into either the myofibroblast or the osteoblast-like phenotype contribute to the cell-mediated disease progression of aortic valve stenosis (AS), resulting in the overall reduced function of the aortic valve and calcification deposition. Microenvironmental cues that mediate the VIC phenotype include, but are not limited to, solubly delivered cytokines and calcifying stimuli, as well as biophysical cues such as extracellular matrix dimensionality and composition. The work presented within this thesis aims to design an in vitro culture platform that recapitulates aspects of the native valve tissue microenvironment, including dimensionality and protein composition, that mineralizes in response to solubly delivered cues, and allows for researchers to answer complex hypotheses behind how AVS progresses in vivo.
With this, we investigated the cell-matrix dynamics and microenvironmental cues which contribute to maladaptive valve repair coordinated by VICs. In Chapter 3, we demonstrate the use of a synthetic, degradable 3D hydrogel which preserves the fibroblast phenotype via the soluble addition of the biochemical cue FGF-2 into the culture medium, or the activation of the myofibroblast via the addition of pro-inflammatory cue TGF-β1. Then, in Chapter 4, we modify the PEG hydrogel platform to form in the presence of a secondary network of collagen type I and develop a system to mediate a mature osteoblast-like VIC phenotype and answer hypotheses regarding calcification inhibitors. Using this PEG + Collagen hydrogel system, we then explore sex differences with respect to mediating mineralization within the hydrogel matrix, using clinical data and human tissues to guide our experimental design in Chapter 5. We discover that osteopontin may be sequestered via collagen into the valve microenvironment to mediate calcification in a sexually dimorphic manner. Finally, in Chapter 6, we further explore the role of FGF-2 in mediating the VIC phenotype to gain a deeper understanding of how this biochemical signaling cue may be contributing to valve repair and extracellular matrix remodeling. Collectively, these results demonstrate the importance of 3D biomaterial platforms to study VIC phenotypic changes and the effects of microenvironmental cues in mediating AVS progression.