SRS297 - Decellularisation barotrauma in rat liver scaffolds compromises scaffold quality and recellularisation capacity

Abstract Liver transplantation remains the only life-saving treatment for end-stage liver failure, but is limited by organ shortage, and transplanted patients face significant immunosuppression-related pathologies. Bioengineering neo-organs by repopulation of decellularised scaffolds with candidate cells offers the possibility of recipient-specific, immunosuppression-free transplantation. Scaffold preparation requires cell removal whilst minimising damage to extra-cellular-matrix (ECM), which provides biophysical/biochemical cellular fate-defining instructions. Portal vein perfusion with decellularisation solution is an established method of generating liver scaffolds, but flowrate effects on scaffold characteristics are poorly defined. We therefore studied effects of decellularisation flowrates on scaffold properties by light-microscopy, immunofluorescence, residual DNA, glycosaminoglycan and hepatocyte growth factor content, structural ECM proteins, and scaffold recellularisation quality with vascular progenitor stem cells. We show that decellularisation solution flowrate is a determining factor of scaffold physical and biochemical quality and report the appearance of previously undescribed disruptions within scaffold ECM with 3D-structure consistent with false passages distinct from vascular lumina. Compared to sub-physiological flowrates (5 mL/min), disruptions were 20-fold more frequent (P = 0.0022) at physiological and higher flowrates (15 and 30 mL/min). Disruptions resulted in poor decellularisation with 5-fold higher levels of remnant DNA (P = 0.0045), out-with established quality criteria. Disruptions were also associated with poor scaffold recellularisation, with 2-fold reduction in sinusoidal repopulation (P = 0.0066), 7-fold imbalance in peripheral to central scaffold cellular engraftment (P = 0.0049), 3-fold and 4-fold reduction in portal and hepatic venous recellularisation efficiency (P = 0.0001). These results demonstrate previously undescribed effects of perfusion decellularisation and contribute to the evolving definition of optimal parameters for the generation of decellularised tissue scaffolds.