| Popis: |
There is a critical need for cartilage regeneration therapies. Not only is cartilage necessary for proper joint function, as deterioration of cartilage leads to osteoarthritis, but it also serves important roles in other places in the body, like in the trachea. Specifically in the articular and tracheal niches, replacement cartilage should have adequate mechanical properties and specific geometries to restore native function. To address these needs, novel strategies to engineer high-density human mesenchymal stem cell (hMSC)-derived cartilage tissues are presented in this dissertation. Bioactive microspheres loaded with chondrogenic transforming growth factor beta 1 (TGF-ß1) were incorporated within some of these tissues for enhanced chondrogenesis. First, scaffold-free cartilage rings and tubes with controlled dimensions were successfully fabricated using custom-made culture wells and a ring-to-tube assembly approach, respectively. The use of TGF-ß1 microspheres in the hMSC rings and tubes significantly improved the quality and quantity of generated cartilage tissue. Next, localized TGF-ß1 presentation within cartilaginous tissues facilitated organized fusion and culture of cartilage tissue building blocks with engineered epithelial and prevascular tissues. Successful development and/or maintenance of tissue-specific phenotypes in this co-culture approach with localized presentation of cues guiding cell differentiation is a promising step toward engineering a functional replacement trachea. Next, extracellular matrix (ECM) scaffolds fabricated from high-density hMSC condensates with and without TGF-ß1 microspheres were shown to support chondrogenesis of re-seeded hMSCs. Importantly, addition of microspheres to hMSC condensates significantly enhanced ECM production and consequently yielded 50% more scaffolds. Additionally, ECM scaffolds were demonstrated to drive chondrogenesis when TGF-ß1 was loaded into them, which suggests improved potential for clinical translatability of this off-the-shelf cartilage regeneration product. Lastly, three different types of bioreactors were designed and engineered or modified for the application of hydrostatic pressure, magnetic bead-induced micromechanical stress and compressive stress to scaffold-free hMSC condensates with the goal to improve the functionality of the engineered cartilage. Stimulation with hydrostatic pressure showed promising evidence of enhanced chondrogenesis in hMSC-derived cartilage, while micromechanical stresses did not improve cartilage tissue formation. Additional studies may further elucidate the impact of each type of stimulus on chondrogenesis. Taken all together, this dissertation developed many strategies and technologies that help advance the field of cartilage tissue engineering. |
