Extracellular vesicles from human liver stem cells reduce injury in an ex vivo normothermic hypoxic rat liver perfusion model

ABSTRACTBackgroundThe gold standard for organ preservation before transplantation is static cold storage, which is unable to fully protect suboptimal livers from ischemia/reperfusion injury. An emerging alternative is normothermic machine perfusion (NMP), which permits organ reconditioning. Here, we aimed to explore the feasibility of a pharmacological intervention on isolated rat livers by using a combination of NMP and human liver stem cells-derived extracellular vesicles (HLSC-EV).MethodsWe established an ex vivo murine model of NMP capable to maintain liver function despite an ongoing hypoxic injury induced by hemodilution. Livers were perfused during 4 hours without (control group, n=10) or with HLSC-EV (treated group, n=9). Bile production was quantified; perfusate samples were collected hourly to measure metabolic (pH, pO2, pCO2) and cytolysis parameters (AST, ALT, LDH). At the end of perfusion, we assessed HLSC-EV engraftment by immunofluorescence, tissue injury by histology, apoptosis by TUNEL assay, and tissue HIF-1α and TGF-β1 RNA expression by quantitative RT-PCR.ResultsDuring hypoxic NMP, livers were able to maintain homeostasis and produce bile. In the treated group, AST (p=0.018) and LDH (p=0.032) levels were significantly lower than those of the control group at 3 hours of perfusion, and AST levels persisted lower at 4 hours (p=0.003). By the end of NMP, HLSC-EV had been uptaken by hepatocytes and EV treatment significantly reduced histological damage (p=0.030), apoptosis (p=0.049), and RNA over-expression of HIF-1α (p

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