Platelet-derived growth factor stabilises vascularisation in collagen-glycosaminoglycan scaffolds in vitro.
Document Type Article
"This is the peer reviewed version of the following article: do Amaral RJFC1, Cavanagh B, O'Brien FJ, Kearney CJ. Platelet-derived growth factor stabilises vascularisation in collagen-glycosaminoglycan scaffolds in vitro. Journal of Tissue Engineering and Regenerative Medicine. 2018., which has been published in final form at https://doi.org/10.1002/term.2789. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving."
Collagen-glycosaminoglycan (CG) scaffolds have been widely developed for a range of regenerative medicine applications. To enhance their efficacy, CG scaffolds have previously been pre-vascularised in vitro using human endothelial cells and mesenchymal stromal cells (hMSC); however, at later time-points, a regression of vascularisation is observed. This is undesirable for longer pre-culture periods (e.g., for partial/full organ regeneration) and for in vitro vascularised tissue model systems (e.g., for drug testing/modelling). We hypothesised that delayed platelet-derived growth factor-BB (PDGF) addition could stabilise vessels, preventing their regression. In 2D, we identified 25ng/ml as a suitable dose that enhanced hMSC metabolic activity and proliferation, without affecting endothelial cells, or migration in either cell type. In our 3D model of CG scaffold vascularisation, early addition of PDGF (day 3), behaved similarly to no PDGF controls. However, PDGF addition at later time-points (i.e., day 4 and 5) with a second addition on day 10, prevented vascular regression. In quantifying our observations, we identified a need for a tool to measure in vitro vascularisation in porous scaffolds. This was a second key objective of this work. A novel ImageJ macro was developed, which allowed us to analyse vessel-like structures, evaluating their number and morphology, and confirmed our qualitative observations. Finally, upregulation of angiogenic genes (ANG1, KDR and TEK2) involved in vessel maturation illustrated how PDGF addition contributed to vascular stability. Taken together, the results suggest that addition of PDGF at specific time-points can be used to stabilise vasculature in CG scaffolds.