Reinforced Collagen Based Scaffolds for Musculoskeletal Tissue Engineering

Articular cartilage has limited self-healing capacity and damaged cartilage can lead to osteoarthritis, and to the need for joint replacement surgery. Biomaterial based scaffolds have shown limited potential to regenerate osteochondral tissue and generally have insufficient mechanical properties to withstand the mechanically demanding and dynamic joint environment. To overcome this limitation, the primary research goal of this thesis was to develop reinforced collagen based scaffolds for musculoskeletal tissue engineering, with focus on articular cartilage, bone, and osteochondral tissue. A process was initially developed in Chapter 2 to produce a range of 3D printed polycaprolactone (PCL) scaffolds with high reproducibility. Optimized 3D printing parameters resulted in a range of PCL scaffolds designs with different compressive moduli and high porosities suitable for bone, cartilage, and osteochondral tissue engineering. A multiple linear regression (MLR) model was developed that can be used to predict the compressive modulus of future designs of 3D printed PCL scaffolds, and to tune it for specific tissues to make suitable composite scaffolds containing different ECM based matrices. In Chapter 3, the PCL scaffold with mechanical properties optimized for cartilage repair 2 was combined with a collagen type I and hyaluronic acid (CI-HyA) matrix, to make a composite scaffold capable of directing MSC differentiation towards chondrogenesis and having mechanical properties suitable for cartilage repair. Sulphated glycosaminoglycan (sGAG) deposition by MSCs undergoing chondrogenic differentiation in the developed composite CI-HyA scaffold led to increased mechanical properties over culture time. In Chapter 4, a PCL scaffold with a high compressive modulus developed in Chapter 2 was combined with a collagen type I and nano-hydroxyapatite (CI-nHA) matrix, to make composite scaffolds with a composition that can promote MSC osteogenic differentiation to regenerate bone. The composite CI-nHA scaffold sustained MSC osteogenic differentiation and achieved an even calcium distribution throughout its 4 whole volume while having a compressive modulus suitable for bone tissue engineering. In Chapter 5, a composite scaffold for osteochondral repair was made by combining the composite scaffolds developed in Chapter 3 and Chapter 4. These bi-layered composite scaffolds were made with a novel one-step freeze drying technique which achieved spatially defined CI-nHA and CI-HyA layers. The bi-layered composite scaffold induced layer-specific MSC differentiation, with sGAG deposition in the cartilage layer which spread to form a sGAG rich matrix over culture time and an increased calcium deposition over culture time in the bone layer, although the results suggested further optimization of the composition of the cartilage layer was needed to minimize risk of calcification. In Chapter 6 a refined cartilage layer consisting of collagen type I, type II and hyaluronic acid was used in a reinforced tri-layered composite scaffold which also contained an intermediate layer consisting of collage type I and hyaluronic acid to reduce off-site calcification in the cartilage layer. The reinforced tri-layered composite scaffold contained a compressive modulus gradient that resisted shear forces and could be securely fixed in the articular joint. The layered matrix composition led to layer-specific MSC differentiation with sGAG rich ECM being deposited throughout the depth of the cartilage layer with calcium deposition contained to the intermediate and bone layers. In conclusion, the thesis has led to the development of a series of composite scaffold systems based on 3D printed PCL designs which incorporated specific collagen-based matrices with biochemical composition tailored to facilitate cartilage, bone, and osteochondral regeneration. The scaffolds have demonstrated potential to facilitate desired MSC differentiation and matrix formation in vitro, and have the mechanical properties required to withstand the compression and shear forces present in articulating joints suggesting can be used as off-the-shelf treatments for regeneration cartilage, bone, or osteochondral defects, with the potential to avoid or delay the need for joint replacement surgery.