RESUMEN
Machine perfusion (MP) allows the maintenance of liver cells in a metabolically active state ex vivo and can potentially revert metabolic perturbations caused by donor warm ischemia, procurement, and static cold storage (SCS). The present preclinical research investigated the metabolic outcome of the MP procedure by analyzing rat liver tissue, bile, and perfusate samples by means of high-field (600 MHz) nuclear magnetic resonance (NMR) spectroscopy. An established rat model of normothermic MP (NMP) was used. Experiments were carried out with the addition of an oxygen carrier (OxC) to the perfusion fluid (OxC-NMP, n = 5) or without (h-NMP, n = 5). Bile and perfusate samples were collected throughout the procedure, while biopsies were only taken at the end of NMP. Two additional groups were: (1) Native, in which tissue or bile specimens were collected from rats in resting conditions; and (2) SCS, in which biopsies were taken from cold-stored livers. Generally, NMP groups showed a distinctive metabolomic signature in all the analyzed biological matrices. In particular, many of the differentially expressed metabolites were involved in mitochondrial biochemical pathways. Succinate, acetate, 3-hydroxybutyrate, creatine, and O-phosphocholine were deeply modulated in ex vivo perfused livers compared to both the Native and SCS groups. These novel results demonstrate a broad modulation of mitochondrial metabolism during NMP that exceeds energy production and redox balance maintenance.
RESUMEN
Hypothermic-oxygenated-machine-perfusion (HOPE) allows assessment/reconditioning of livers procured from high-risk donors before transplantation. Graft referral to HOPE mostly depends on surgeons' subjective judgment, as objective criteria are still insufficient. We investigated whether analysis of effluent fluids collected upon organ flush during static-cold-storage can improve selection criteria for HOPE utilization. Effluents were analyzed to determine cytolysis enzymes, metabolites, inflammation-related mediators, and damage-associated-molecular-patterns. Molecular profiles were assessed by unsupervised cluster analysis. Differences between "machine perfusion (MP)-yes" vs. "MP-no"; "brain-death (DBD) vs. donation-after-circulatory-death (DCD)"; "early-allograft-dysfunction (EAD)-yes" vs. "EAD-no" groups, as well as correlation between effluent variables and transplantation outcome, were investigated. Livers assigned to HOPE (n = 18) showed a different molecular profile relative to grafts transplanted without this procedure (n = 21, p = 0.021). Increases in the inflammatory mediators PTX3 (p = 0.048), CXCL8/IL-8 (p = 0.017), TNF-α (p = 0.038), and ANGPTL4 (p = 0.010) were observed, whereas the anti-inflammatory cytokine IL-10 was reduced (p = 0.007). Peculiar inflammation, cell death, and coagulation signatures were observed in fluids collected from DCD livers compared to those from DBD grafts. AST (p = 0.034), ALT (p = 0.047), and LDH (p = 0.047) were higher in the "EAD-yes" compared to the "EAD-no" group. Cytolysis markers and hyaluronan correlated with recipient creatinine, AST, and ICU stay. The study demonstrates that effluent molecular analysis can provide directions about the use of HOPE.
