The major aim of this project was to develop a biologically active bone scaffold that could induce vascularization in critical-size defects (CSD) and hence bone formation. In this study, functionalization of three-dimensional (3D) printed biphasic calcium phosphate (BCP) scaffolds was investigated. The first functionalization approach involved printing scaffolds with two different pore geometries and sizes; square (400μ) and round (800μ). The second was by coating scaffolds with DAR16-II; a self-assembly peptide that forms a hydrogel nanostructure mimicking extracellular matrix (ECM). A rabbit model was used to study these functionalization methods; square and round pore scaffolds with and without DAR16-II coating were implanted into experimental rabbit calvaria bone CSD defects. After 8 weeks, animals were killed and tissue was processed for histomorphometric analysis. Histological evaluation showed that bone formation was pore size and geometry independent while DAR16-II was successful in inducing bone formation compared to non-coated scaffolds. The following in vitro studies aimed towards understanding the basic cell response that enhanced bone formation in vivo. Human mesenchymal stem cells (MSCs) were used to identify the osteogenic potential of DAR16-II. Molecular analysis and mineralization staining showed that DAR16-II lacks osteoinductive properties. However, DAR16-II preserved cell viability when used as a BCP coating in vitro. In addition, DAR16-II exhibited angiogenic potential upon culturing with human umbilical vein endothelial cells (HUVECs) in vitro. DAR16-II induced the spreading of endothelial cells, activation and tubular-structure formation. Angiogenesis Real time-2 (RT2) polymerase chain reaction (PCR) array was used for gene expression analysis and showed that DAR16-II angiogenic effect was regulated by overexpression of endoglin (ENG or CD105), a clade E member of the serine protease inhibitor-1 (SERPIN-1) and β-Actin (ACTB) and down-regulation of VEGF receptor I (Flt1) and VEGF receptor II (KDR) Flt1. Furthermore, DAR16-II enhanced attachment of monocyte THP-1 cells. Results have demonstrated that DAR16-II add a proactive factor to BCP scaffolds. The proposed functionalization methodology increases the potential of enhancing vascularization and bone formation within ceramic scaffolds.
Synergizing angiogenesis and osteogenesis in a smart bone substitute
Alfayez, E. S. (Author). 2016
Student thesis: Doctoral Thesis › Doctor of Philosophy