3D Bioprinting of Anatomically Accurate Implants for Meniscus Tissue Engineering

Abstract

Menisci are soft tissues essential for load bearing and stress distribution in the knee joint. Meniscal injuries are common in all age groups and can lead to degeneration of the joint. While currently available surgical debridement, meniscal repair and replacement strategies provide short-term relief, the ideal solution would be to facilitate meniscus regeneration. This has motivated increased interest in meniscus tissue engineering strategies to create more advanced cell-based implants for joint preservation. The overall aim of this thesis was to 3D bioprint a cell-laden and anatomically accurate engineered meniscus construct as an implant to replace meniscus tissue after a total or partial meniscectomy. Realizing this aim required the development of polymer scaffolds and bioinks with specific biomechanical properties and the potential to support fibro-cartilaginous tissue deposition by encapsulated cells. To produce scaffolds with biomechanical properties mimetic of the native meniscus, fused deposition modelling (FDM) was first used to 3D print polycaprolactone (PCL) fibre networks with varying fibre diameter, spacing and print patterns. Using this approach it was possible to produce porous PCL scaffolds with axial compressive properties and radial tensile properties similar to human meniscus tissue. Next, a composite construct was developed which consisted of a mesenchymal stem cell (MSC) laden interpenetrating network (IPN) hydrogel based on alginate and gelatin methacryloyl (GelMA), reinforced with a 3D printed PCL fibre network. In a rheological analysis the developed hydrogel showed shear thinning properties making it suitable for putative 3D bioprinting applications. Furthermore the biomaterial supported robust chondrogenesis of MSCs in vitro, with the engineered tissue displaying dynamic biomechanical behavior similar to bi-phasic soft tissues like the meniscus. Recognizing that the meniscus is spatially heterogenous, with the inner zone compositionally distinct to the outer zone, this thesis next sought to functionalize the alginate and GelMA IPNs with solubilized extracellular matrix (ECM) isolated from the inner and outer zones of porcine menisci. While the addition of ECMs improved the shear thinning properties of the developed hydrogel bioinks, it did not dramatically alter the capacity of the inks to support the fibrochondrogenic differentiation of MSCs. Finally, a novel 3D bioprinting technique termed "z-printing" was implemented to facilitate the relatively rapid printing of complex, large scale tissue engineered constructs. Using this approach a meniscus construct with a size and both internal and external structure similar to that of the human meniscus was produced, using previously optimized printing parameters. Composite constructs with circumferential and radial fibre orientations and viscoelastic biomechanical properties were produced which mimicked anisotropic structure and heterogeneous properties of native meniscus tissue. To conclude, this thesis describes the development of a 3D bioprinted meniscus construct which mimics the external size and shape of the human meniscus, as well as aspects of its internal heterogeneous structure. In the future such 3D bioprinted implants could provide the basis of a new treatment strategy for meniscal injuries.