Engineering Gelatin-Based Biomatrices for Pre-vascularisation of Bone Analogues

Abstract

The current treatment to replace a large missing bone segment is by transplantation with autologous tissue. Such reconstructive surgeries are accompanied by long operation times and donor site morbidity. To overcome these shortcomings in future medicine and to preserve, restore or improve the functions of the affected tissue of a patient, the technology to engineer tissues in the laboratory is arising. In this thesis, pre-existing gelatin-based hydrogels were evaluated regarding their suitability as matrices for the engineering of vascularised bone. Moreover, a gelatin- polyethylene glycol (PEG)- based hydrogel system (gelPEG) was newly developed to address the shortcomings of available gelatin-based hydrogel platforms. Gelatin-methacryloyl (gelMA) and gelatin-transglutaminase (gelTG) hydrogels were limited for the intended application to fabricate soft hydrogels with low polymer concentrations and/or crosslinking density. With such a hydrogel design, these pure gelatin-based materials were physically instable over culture time with stem cells. However, the newly developed gelPEG matrix material proved to be adequately soft with a suitable speed of cell-mediated degradation, which allows for differentiation towards the vasculogenic and osteogenic lineages. Moreover, the developed gelPEG platform could also be applied for other tissue culture techniques, such as for liver organoids, which are currently done in Matrigel, a mouse tumour-based matrix. Importantly, this novel tailorable hydrogel is easily customised with tissue-specific cues that might prove valuable for other tissue engineering approaches that combine pre-vascularisation approaches alongside specific tissue development. In this thesis, endothelial colony forming cells (ECFCs) were used which were obtained from umbilical cord blood and combined in a co-culture with multipotent mesenchymal stromal cells (MSCs), obtained from bone marrow. By using these cells, for the first time, the use of clinically relevant stem cells for the generation of complex multiscale vascularised bone-like tissue constructs was demonstrated. The multiscale vasculature existed of a central channel in the hydrogel construct, which was surrounded by a capillary-like network. The next steps will lie in perfusion of such constructs and in vivo models to explore questions, such as how much predeveloped complexity of such tissue engineered constructs is needed to serve the goal to repair and restore the function of damaged tissue. Platforms, such as in this thesis developed gelPEG system, might serve as a stepping stone to bridge the gap between tissue engineering at the laboratory bench and clinically relevant in vivo models.