Endochondral bone regeneration: Stepping stones towards clinical translation

Abstract

Bone possesses a great regenerative capacity, as the majority of injuries reach resolution through complete healing. Nevertheless, incomplete healing is observed in 10% of all fractures, usually because the size of the defect exceeds the intrinsic healing capability of the tissue. These non-healing defects could be the result of cancer resection, radiation therapy, trauma, infection, or congenital abnormalities. Autograft, the transplantation from patient’s own bone isolated from a different location (e.g. fibula) is currently considered the golden standard. However, this procedure presents several disadvantages including a high risk of complications such as infection, nerve injury and donor site morbidity. Due to the limitation of the currently available treatments, efforts in the field of regenerative medicine are focused on engineering a bone substitute by combining relevant cell types with biomaterials and bioactive stimuli. A promising strategy to induce bone regeneration exploits the chondrogenic potential of mesenchymal stem cells (MSCs) to mimic the endochondral ossification process. To mimic this process, a cartilaginous soft callus can be engineered in vitro and implanted at the bone defect site. Once implanted, the chondrocytes will acquire a hypertrophic phenotype, the engineered cartilage will be invaded by blood vessels, host osteoblasts and osteoclasts, and eventually it will be converted into bone. The feasibility of recapitulating the endochondral process for regenerative purposes has been widely established in the last 15 years. Nevertheless, several challenges still need to be faced to transform these scientific results into a realistic therapeutic option. In particular, numerous aspects that pertain to the in vitro development of the engineered cartilage need to be standardized to obtain a predictable regenerative effect. For example, the high variability and unpredictability of the chondrogenic potential of MSCs isolated from different patients will preclude this treatment to patients whose MSCs present limited chondrogenic capacity. Therefore, the overall aim of this thesis is to improve the clinical translation of endochondral bone regeneration (EBR)-based strategies to treat critical-sized bone defects. To achieve this goal, the first part of this thesis was focused on investigating cell-based and biomaterial-based strategies to engineer a cartilaginous template with predictable features in vitro. To overcome the variability associated with the use of bone marrow-derived MSCs, in Chapter 2, the feasibility of using dental pulp stem cells as an alternative cell source was explored. In addition, in Chapters 3 and 4, the possibility of enhancing the chondrogenic differentiation of MSCs by exploiting the characteristics of vitreous humor (VH) as supportive material was evaluated. In the second part of this thesis we focused on improving the clinical translatability of EBR- based approaches by exploring the possibility of using allogeneic MSCs to engineer the cartilaginous template. Thus, in Chapter 5 and 6 the immune response triggered by the implantation of non-autologous, chondrogenically differentiated MSCs and its influence on the regeneration of bone defects was investigated. Finally, aiming at developing an off-the-shelf product, in Chapter 7, the feasibility of using allogeneic, devitalized cartilaginous implants as a stand-alone treatment for the regeneration of critical size bone defects was explored.