Translational challenges in bone tissue engineering

Abstract

The research performed for this thesis focused at strategies to improve bone graft substitutes for future clinical applicability. We started by investigating the value of cell based tissue engineered constructs. First we showed that at the ectopic location, bone formation was only present when BLI signal was present until the end of the implantation period, indicating that living MSCs were necessary for bone formation. By immunohistochemistry we showed, that the implanted MSCs were present in the newly formed bone as bone cell. For the spinal fusion location the question whether the implanted MSCs contribute to the bone regeneration remained unanswered. Next, we created hybrid constructs by using allogeneic MSCs as off-the-shelf components, and showed their effectivity in goat models both at the ectopic as well as the orthotopic location. Before allogeneic MSCs can be used in clinical practice further studies need to be performed to analyze safety in man. In other studies performed in goats we found that location has great influence on the effectiveness of MSCs. They show an additive value in enhancing bone formation at the ectopic location, e.g. in rats, but at the orthotopic location this beneficial effect is only present during the first few weeks, and not after implantation of 3-4 months. From this we can conclude that MSC seeding is not necessary for bone formation at clinically relevant orthotopic locations. Furthermore, early EPCs were coseeded with MSC to stimulate vasculogenesis, which is a prerequisite for osteogenesis. We found that in spite of the positive effect that both cell types (MSCs and EPCs) exerted on proliferation of the other in vitro, co-seeding of MSCs and EPCs did not improve bone formation. Bone-marrow-derived EPCs alone did reveal enhanced bone formation as compared to control implants without cells. The second aspect of this thesis emphasized on construct optimization, involving material improvement and investigation of bioactives. PLG, containing growth factors, was used in three studies in combination with (allogeneic) MSCs and EPCs to enhance bone formation. Results were highly variable, which was also reported in animal and human studies in the literature, making PLG in our current opinion an unreliable component for bone tissue engineering purposes. Next, the value of timing of sequentially released BMP-2 and VEGF was analyzed at ectopic and orthotopic sites in dogs. A positive effect of growth factors on bone formation was seen at both locations, which is in agreement with reports from literature. With respect to timing of the growth factors, we were the first to show that a differential timing of dual VEGF and BMP-2 delivery did not influence orthotopic bone regeneration, whereas at the ectopic site early release of BMP-2 significantly enhanced bone formation. Finally we performed an instrumented posterolateral fusion, using a new TCP, and demonstrated that the cell-free TCP constructs and MSC-seeded TCP implants performed not different from autograft, implying a future role for osteoinductive materials such as the TCP for spinal fusion. From preclinical studies we conclude that cell seeding is almost never needed in bone regeneration.