SYNERGISTIC INFLUENCES OF OSCILLATING PRESSURE, PERFUSION, AND GROWTH FACTORS ON CHONDROGENESIS IN A NOVEL CENTRIFUGAL BIOREACTOR
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Defects in articular cartilage (AC) cannot be healed spontaneously primarily because AC is avascular and aneural. Currently available surgical techniques are also incapable of restoring damaged AC. Therefore, AC tissue engineering (ACTE) has begun to attract much attention as an alternative therapy. This dissertation is focused on identifying optimal methods to engineer AC, including finding the best cell source, formulating an ideal biochemical cocktail, designing a bioreactor that mimics the mechanical forces that AC experiences in vivo, and using a biocompatible scaffold to provide the needed structural support for the engineered constructs. First, a comprehensive review was conducted to systematically weigh the advantages and disadvantages associated with cell types that are commonly used in ACTE, including articular chondrocytes (AChs), non-AChs, and mesenchymal stem cells (MSCs). A unique approach was also considered that involves co-culturing AChs with MSCs to overcome the drawbacks associated with using each of these cell types in isolation. Based on this analysis, comprehensive cell-type and culture-condition recommendations were made available for future studies. The contents of that review formed the basis for our subsequent efforts to define a “gold standard” growth-medium cocktail that can best promote chondrogenesis, and to develop insight into temporal effects and growth-factor adjustments during the culture period. Finally, the most effective conditions for engineering AC cannot be achieved without incorporating into the growth environment mechanical forces similar to those that AC encounters continuously. Currently available bioreactors are not capable of simultaneously applying shear stress and oscillating hydrostatic pressure (OHP), which are both important physical forces encountered by AC in vivo. We designed a new type of perfusion/pressurized bioreactor that could accomplish this goal. Using this bioreactor, we were able to show that TGF-β3 supplementation and OHP can work synergistically to improve the mechanical properties of tissues engineered from adipose-derived stem cells. To further improve the properties of the engineered tissue, we encapsulated cells in agarose hydrogels and showed that such encapsulation significantly improves extracellular matrix synthesis. For the first time, our results indicate that shear stress and OHP can be used to synergistically enhance the glycosaminoglycogen and collagen contents of engineered AC tissue.