Co-Culture of Precision-Cut Liver Slices and Liver-Derived Mesenchymal Stromal Cells

Abstract

Due to the limited availability of suitable livers for transplantation and the increasing use of suboptimal donor livers, which are more prone to ischemia-reperfusion injury (IRI), there is an urgent need for novel graft-ameliorating therapies. The promising capacities of (liver-derived) mesenchymal stromal cells ((L-)MSC) in mitigating IRI and their integration with hypothermic oxygenated machine perfusion (HOPE) present a novel paradigm for augmenting graft quality. However, still little research exists into the effects on the liver of (L-)MSC administered during HOPE. Moreover, human livers for research are scarce and, therefore, studies to show safety and efficacy are challenging. To address these challenges, this study investigated the use of porcine precision-cut liver slices (PCLS) as a tool to model the administration of MSC during HOPE. First, optimisation of PCLS procurement and culture was undertaken, exploring methods such as the use of well inserts and different culture media to enhance the retained structural integrity of the slices. Subsequently, a 24-hour co-culture experiment of porcine PCLS (n=3 livers, n=135 PCLS) and L-MSC was conducted, where L-MSC were added at the beginning of a two-hour hypothermic oxygenated culture. The findings of this experiment demonstrated the ability of MSC to adhere to the liver tissue during the two hours in a hypothermic oxygenated environment. Additionally, the effect of MSC on PCLS viability, morphology, cell death, regeneration, liver injury markers, functionality, and gene expression profiles was investigated. While no significant differences between co-cultured and control PCLS were observed, there was a clear trend towards increased inflammation, cell damage, and cell death in the co-cultured slices. Overall, this research contributes to the understanding of the potential implications of (L-)MSC administration during hypothermic oxygenated perfusion and highlights the utility of PCLS as an intermediate experimental model for studying transplant-related IRI.