Speaker
Description
"The establishment of vascularisation is one of the current challenges, especially in critically sized bone defects [1,2]. Therefore, the aim of the present work is to develop a biomimetic hydrogel scaffold mimicking the bone osteoid in which proper vasculature is induced in order to maximally stimulate osteogenic regeneration. Thiol-ene step-growth photocrosslinking was herein selected to enable superior control, homogeneous network formation and lower radical concentrations compared to traditional chain-growth systems [3]. The networks were benchmarked against traditional chain-growth systems in a physico-chemical and an in vitro osteo- and angiogenic evaluation.
For osteogenic purposes, a novel norbornene-modified aminated gelatin was developed with a degree of substitution of 135% compared to the amount of amines present in native gelatin type B (by addition of only 1.2 equivalents of norbornene carboxylic acid). It is, to the best of our knowledge, the highest substitution reported for norbornene-functionalised gelatins [4]. Thiol-ene crosslinking with thiolated gelatin as cell-interactive crosslinker resulted in networks with full norbornene conversion and a gel fraction of 99%. The increase in storage modulus compared to non-aminated thiol-ene and gelatin-methacryloyl systems (through reaction with the same amount of equivalents) was attributed to an increase in chemical crosslinking (1.8-fold and 4.6-fold respectively) and a decrease of swelling (3.2-fold and 2.9-fold). The biodegradation properties of this hydrogel were preserved and direct contact cell viability data indicated an extended morphology and excellent biocompatibility (96% viability after 7 days). Osteogenic differentiation seeding tests indicated an increase in alkaline phosphatase (ALP) production after 7 days (2.8-fold and 1.5-fold) and a rise in calcium deposition after 21 days (5.2-fold and 1.7-fold). In addition, the incorporation of amorphous calcium phosphate into the thiol-ene hydrogel further contributed to the osteogenicity (1.1-fold increase in ALP-production and 1.2-fold increase in stem cell-based calcium deposits).
Thiol-ene systems were also benchmarked against chain-growth systems with regard to angiogenic stimulation. The homogeneous networks showed similar attachment, yet a more pronounced vascular signalling capacity in terms of fibroblast growth factor-1 (1.7-fold increase after 1 day) and epidermal growth factor (1.2-fold increase after 1 day) secretion compared to the more heterogeneous chain-growth networks which signalled mainly through the production of placental growth factor (2-fold increase after 1 day). Moreover, the thiol-ene networks showed an increased seeded endothelial cell viability (1.2-fold after 7 days) and metabolic activity (1.3-fold after 7 days). The incorporation of human placenta substrate to the thiol-ene networks greatly stimulated the sprout junction density, the total tube length and the number of branches within the developed vascular network in these hydrogels.
The results show that highly controlled networks could be created with tailored topology and viscoelastic behaviour which can stimulate angio- and osteogenesis. Ongoing work focusses on the processing of these hydrogels as bio-inks through extrusion-based 3D-printing and evaluation of the 3D-printed constructs towards vascularised osteogenesis.
[1] Parmentier L, et al. Materials, 2020;
[2] Nguyen LH, et al. Tissue Eng Part B Rev, 2012;
[3] Hoyle C, Bowman C. Angew Chem Int Ed, 2010;
[4] Van Hoorick, J, et al., Macromol Rapid Commun, 2018."
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