Assessment of Mitochondrial Respiration During Hypothermic Storage of Liver Biopsies Following Normothermic Machine Perfusion.

Organ quality can be assessed prior to transplantation, during normothermic machine perfusion (NMP) of the liver. Evaluation of mitochondrial function by high-resolution respirometry (HRR) may serve as a viability assessment concept in this setting. Freshly collected tissue is considered as optimal sample for HRR, but due to technical and personnel requirements, more flexible and schedulable measurements are needed. However, the impact of cold storage following NMP before processing biopsy samples for mitochondrial analysis remains unknown. We aimed at establishing an appropriate storage protocol of liver biopsies for HRR. Wedge biopsies of 5 human livers during NMP were obtained and assessed by HRR. Analysis was performed after 0, 4, 8, and 12 h of hypothermic storage (HTS) in HTK organ preservation solution at 4°C. With HTS up to 4 h, mitochondrial performance did not decrease in HTS samples compared with 0 h (OXPHOS, 44.62 [34.75-60.15] pmol·s-1·mg wet mass-1 vs. 43.73 [40.69-57.71], median [IQR], p > 0.999). However, at HTS beyond 4 h, mitochondrial respiration decreased. We conclude that HTS can be safely applied for extending the biopsy measurement window for up to 4 h to determine organ quality, but also that human liver respiration degrades beyond 4 h HTS following NMP.

The BAR Score Predicts and Stratifies Outcomes Following Liver Retransplantation: Insights From a Retrospective Cohort Study.

Liver retransplantation (reLT) yields poorer outcomes than primary liver transplantation, necessitating careful patient selection to avoid futile reLT. We conducted a retrospective analysis to assess reLT outcomes and identify associated risk factors. All adult patients who underwent a first reLT at the Medical University of Innsbruck from 2000 to 2021 (N = 111) were included. Graft- and patient survival were assessed via Kaplan-Meier plots and log-rank tests. Uni- and multivariate analyses were performed to identify independent predictors of graft loss. Five-year graft- and patient survival rates were 64.9% and 67.6%, respectively. The balance of risk (BAR) score was found to correlate with and be predictive of graft loss and patient death. The BAR score also predicted sepsis (AUC 0.676) and major complications (AUC 0.720). Multivariate Cox regression analysis identified sepsis [HR 5.179 (95% CI 2.575-10.417), p < 0.001] as the most significant independent risk factor for graft loss. At a cutoff of 18 points, the 5 year graft survival rate fell below 50%. The BAR score, a simple and easy to use score available at the time of organ acceptance, predicts and stratifies clinically relevant outcomes following reLT and may aid in clinical decision-making.

Bioenergetic and Cytokine Profiling May Help to Rescue More DCD Livers for Transplantation.

The majority of organs used for liver transplantation come from brain-dead donors (DBD). In order to overcome the organ shortage, increasingly donation after circulatory death (DCD) organs are also considered. Since normothermic machine perfusion (NMP) restores metabolic activity and allows for in-depth assessment of organ quality and function prior to transplantation, such organs may benefit from NMP. We herein compare the bioenergetic performance through a comprehensive evaluation of mitochondria by high-resolution respirometry in tissue biopsies and the inflammatory response in DBD and DCD livers during NMP. While livers were indistinguishable by perfusate biomarker assessment and histology, our findings revealed a greater impairment of mitochondrial function in DCD livers after static cold storage compared to DBD livers. During subsequent NMPs, DCD organs recovered and eventually showed a similar performance as DBD livers. Cytokine expression analysis showed no differences in the early phase of NMP, while towards the end of NMP, significantly elevated levels of IL-1ß, IL-5 and IL-6 were found in the perfusate of DCD livers. Based on our results, we find it worthwhile to reconsider more DCD organs for transplantation to further extend the donor pool. Therefore, donor organ quality criteria must be developed, which may include an assessment of bioenergetic function and cytokine quantification.

Mitochondrial respiration during normothermic liver machine perfusion predicts clinical outcome.

BACKGROUND: Reliable biomarkers for organ quality assessment during normothermic machine perfusion (NMP) are desired. ATP (adenosine triphosphate) production by oxidative phosphorylation plays a crucial role in the bioenergetic homeostasis of the liver. Thus, detailed analysis of the aerobic mitochondrial performance may serve as predictive tool towards the outcome after liver transplantation. METHODS: In a prospective clinical trial, 50 livers were subjected to NMP (OrganOx Metra) for up to 24.ßh. Biopsy and perfusate samples were collected at the end of cold storage, at 1.ßh, 6.ßh, end of NMP, and 1.ßh after reperfusion. Mitochondrial function and integrity were characterized by high-resolution respirometry (HRR), AMP, ADP, ATP and glutamate dehydrogenase analysis and correlated with the clinical outcome (L-GrAFT score). Real-time confocal microscopy was performed to assess tissue viability. Structural damage was investigated by histology, immunohistochemistry and transmission electron microscopy. FINDINGS: A considerable variability in tissue viability and mitochondrial respiration between individual livers at the end of cold storage was observed. During NMP, mitochondrial respiration with succinate and tissue viability remained stable. In the multivariate analysis of the 35 transplanted livers (15 were discarded), area under the curve (AUC) of LEAK respiration, cytochrome c control efficiency (mitochondrial outer membrane damage), and efficacy of the mitochondrial ATP production during the first 6.ßh of NMP correlated with L-GrAFT. INTERPRETATIONS: Bioenergetic competence during NMP plays a pivotal role in addition to tissue injury markers. The AUC for markers of outer mitochondrial membrane damage, ATP synthesis efficiency and dissipative respiration (LEAK) predict the clinical outcome upon liver transplantation. FUNDING: This study was funded by a Grant from the In Memoriam Dr. Gabriel Salzner Stiftung awarded to SS and the Tiroler Wissenschaftsfond granted to TH.