Oral Presentation 30th Australian and New Zealand Bone and Mineral Society Annual Scientific Meeting 2020

Engineering the vasculature for bone engineering (#39)

Khoon S Lim 1
  1. Department of Othopaedic Surgery & Musculoskeletal Medicine, University of Otago Christchurch, New Zealand.

Bone grafting remains as one of the most common surgical procedure to treat fracture bones and bone injury, with over two million grafting procedures performed annually worldwide. To date, synthetic bone grafts are commonly used clinically to treat these bone injuries. Although these bone grafts have been previously validated and proven to support bone growth, the maintenance of their long-term survival remains a challenge. This phenomenon might be due to the lack of vascularization, where the slow and limited angiogenic ingrowth from the host tissue is insufficient to provide nutrients and oxygen to the newly formed bone. This might also subsequently affect the degree of osseointegration with the surrounding native bone. Therefore, there is a need to develop novel strategies to improve the vasculogenic capacity of these synthetic bone grafts. In this study, we evaluated the vasculogenesis and biofabrication potential of a photo-polymerisable thiol-ene gelatin based hydrogel. Gelatin (10wt%) was reacted with carbic anhydride (20wt%), at 50˚C for 24h with pH kept in the range of 7.5 – 8 to produce gelatin-norbornene (GelNor). 5wt% GelNor hydrogels were photo-polymerised (400-450nm, 30mW/cm2, 3min, Ruthenium (Ru)/Sodium Persulphate (SPS) as photoinitiators) with thiolated molecules as crosslinking agents. The physico-mechanical properties were characterised with varying crosslinking parameters (Nor/thiol ratio, Ru/SPS concentration and crosslinking agent). Human umbilical vein endothelial cells (HUVEC) were co-encapsulated within GelNor hydrogels with human mesenchymal stromal cells (MSCs). Co-cultures in casted hydrogel disks (Ø5x1mm) were maintained in endothelial growth media for 7 and 14 days, following fixation and immunohistochemical evaluation (CD31/F-actin). Scaffolds with interconnected channels were fabricated by casting GelNOR macromer over 3D plotted Pluronic127© which served as a sacrificial template. GelNOR was successfully synthesised with a 45% degree of modification. Varying Nor:SH (DTT) ratios, photoinitiator concentrations and crosslinking agents resulted in tailorable sol fractions, mass swelling ratios and compression modulus. HUVECs were co-encapsulated with MSCs in gelNOR hydrogels, showing high viability (>90%) and a retained HUVEC phenotype over the cell-culture period. These conditions were able to facilitate the formation and stabilisation of interconnected vessel-like structures. Interconnected channels were successfully generated. In conclusion, we have shown that GelNOR hydrogels, with tailorable physico-chemical properties, can be used to promote in vitro vasculogenesis for large biofabricated bone grafts.