Coupling normothermic machine perfusion with mesenchymal stem cell bioreactors: benefits exerted on the rat liver

Abstract

Abstract Administration of mesenchymal-stem-cells (MSCs) during Normothermic-Machine-Perfusion (NMP) reduces ischemia/reperfusion injury, but the strategy to apply cell-based therapies ex-situ can hamper their effectiveness. To fully harness MSCs potential, we developed an advanced perfusion system equipped with a MSCs-bioreactor and investigated its benefits on the rat liver. The study involved two work packages: 1) Development: bioreactors seeded with 2x107 human adipose tissue-derived MSCs were connected to the NMP circuit and subjected to 4h-liverless perfusion (Liverless-NMP,n=5); 2) MSCs-bioreactor-based liver NMP: rat livers were perfused for 4h on the NMP circuit coupled with the MSCs-bioreactor (NMP+bioreactor,n=5) or without it (NMP,n=5). MSCs remained metabolically active during liverless-NMP and showed a preserved expression of stemness-related markers. When used for liver NMP, MSCs changed their secretome in a liver-tailored manner. Organs exposed to the MSCs-bioreactor-based perfusion produced more bile, released less damage biomarkers, and showed higher ATP content with reduced succinate accumulation than those subjected to NMP alone. A lower release of pro-inflammatory mediators and the induction of factors involved in resolution/regeneration were also observed. This proof-of-concept study provides a novel MSCs-based protocol for liver NMP, thereby creating a modern perfusion platform able to preserve mitochondrial function, sustain hepatic cell viability, reduce inflammation, and trigger healing processes.