Printing bone : the application of 3D fiber deposition for bone tissue engineering

Abstract

Bone chips are used by orthopaedic surgeons for treating spinal trauma and to augment large bone defects. A potential alternative to autologous bone is regeneration of bone tissue in the lab by developing hybrid implants consisting of osteogenic (stem) cells seeded on supportive matrices. Application of large bone grafts in the operation room is not a clinical reality yet due to, among other factors, cell death in de core of cm-scale implants. One of the reasons for this is the time needed for vascularisation of the grafts after in vivo implantation. Also, current hybrid grafts are rather simplistic often comprising of one cell type seeded on one type of a biomatrix, while native bone tissue consists of multiple cells, matrix molecules and growth factors each with own spatiotemporal pattern of presentation. For development of functional, large-size grafts it would be attractive to employ techniques that approach the complex organization of bone tissue. Organ- or tissue printing is a novel approach of regenerative medicine, which enables mimicking anatomical organization of tissues by developing 3D-structured cell-laden multimaterial scaffolds. In this thesis we studied the application of 3D fiber deposition (an organ printing technique) for development of (vascularized) bone grafts. We first present a literature overview addressing studies that describe a positive effect biomimicking approaches the functionality of tissue-engineered grafts, and define parameters necessary for printing of cell-laden constructs in their application as bone grafts. Hydrogel matrices are highly hydrated polymer networks used as scaffolding materials in organ printing. Of all synthetic hydrogels, photopolymerizable hydrogels formed by UV-exposure of photosensitive polymers in the presence of photoinitiators, are currently one of the most promising materials for skeletal TE due to their mechanical stability. The influence of photoexposure on cell-cycle progression, which is often cautioned for, is well tolerated by encapsulated cells in hydrogels. One of the other principal printing components is the 3D fiber deposition machine, and we tested its applicability to print cell-laden hydrogels, demonstrating that osteogenic progenitors and hydrogels can be deposited simultaneously. We showed that osteogenic progenitors survive the deposition process and retain the ability to differentiate after printing. Based on our findings we conclude that 3DF is a well-suited tool for printing of viable osteogenic grafts. Tailoring of dispensing parameters such as fiber distance, orientation, pressure, and speed was employed to modulate the characteristics of the printed scaffolds including their porosity and mechanical properties. Porous scaffold design is vital for functionality of embedded cells in vitro and supports tissue development in vivo, while addition of calcium phosphate microparticles is useful to further enhance bone formation. The second part of this thesis addressed the development of multicellular grafts whereby heterogeneous grafts demonstrate retention of cell organization introduced by the printing and result in heterogeneous matrix formation, both for vascularised bone grafts and osteochondral implants. Cumulatively, these results demonstrate the possibility of manufacturing viable and functional heterogeneous tissue constructs by a 3D fiber deposition technique, which could potentially be used for the repair of bone defects and osteochondral lesions.