Conference Agenda

Mitochondrial transplantation (MTx) is a promising new therapeutic where healthy donor mitochondria are injected into a host following acute ischemic injury, such as cardiac arrest (CA) or ischemic stroke. Across three mammalian models, our lab has shown the effectiveness of MTx in treating ischemic injury following CA. Our model induces severe CA in rats, mice, and pigs, followed by resuscitation. In control groups, up to 90% of animals died within 72 hours of resuscitation, which mimics the epidemiology of CA in humans. However, when fully functional endogenous mitochondria were administered via intravenous infusion to animals immediately following resuscitation, we found significant improvements in multiple metabolic indices of ischemic damage like faster lactate clearance, and in functional outcomes like improved neurological recovery and higher survival rates.

How mitochondrial transplantation protects the brain and heart in this model is currently unknown. However, our data suggest two core hypotheses: first, a two-phase mitochondrial trafficking model where a rapid, immunomodulated uptake phase in the bloodstream is followed by a delayed relocalization to the tissues most affected by ischemia. Second, the data indicate that a critical-for-survival but poorly described dose-response for MTx exists. Our most recent results suggest that the laboratories around the world exploring MTx, including those involved with ongoing human trials, may be using a “less than optimal” dosing for ongoing studies.

This talk will review the current state of MTx human subject research, discuss our current data in the context of this research, and suggest critical inflection points for the future of translational research in mitochondrial medicine as a novel approach to ischemic injury.

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Ischemia-reperfusion (IR) injury remains a major contributor to organ dysfunction following transient hypoxic insults. Although numerous interventions have been found effective to reduce IR injury in preclinical models, none of these therapies have been successfully translated to the clinical setting. In the context of the persistent translational gap, we systematically investigated the mechanisms implicated in IR injury using kidney donation and transplantation as a clinical model of IR.

Whilst our results do not implicate traditional culprits such as reactive oxygen species, complement activation or inflammation as triggers of IR injury, they reveal a clear metabolic signature for renal IR injury. This discriminatory signature of IR injury is consistent with a post-reperfusion metabolic paralysis caused by a Krebs cycle defect at the level of the oxoglutarate complex resulting in state of normoxic glycolysis. Concomittant involves high-energy phosphate depletion, glycolysis and a compensatory activation of all catabolic routes may result in vicious cycle of progressive reductive stress. Against this background, the picture emerges that clinical IR injury is driven by reductive stress rather than oxidative stress..

Purpose:

Currently used markers to assess liver viability and predict biliary complications (i.e., bile flow, biliary chemistry) lack validation and robustness in large transplant cohorts. This study validates current parameters used to predict biliary complications during normothermic machine perfusion (NMP) and explores the predictive value of Flavin-mononucleotide (FMN)-levels in bile.

Methodology:

All transplants with normothermic machine perfusion (NMP, OrganOx-Metra®) performed at our center were included. Donor and recipient risk factors were analyzed together with biliary complications and required interventions. ROC-curve analyses were performed for overall and severe biliary complications based on the area-under-the-curve (AUC) of currently applied viability parameters (i.e., bile flow, pH, HCO^3-, Glucose), and bile spectroscopy for Flavin-mononucleotide (FMN) levels.

Results:

Between 10/2022 and 11/2023, 200 livers underwent endischemic NMP, resulting in 188 liver transplantations (DBD:132; DCD:56) with 150 recipients achieving a follow-up of ≥3months.

Outcomes were good with a low number of 15.2% overall biliary complications with a first diagnosis after a median of 33days (IQR:11.5; 47.3). A median of 2 stents were needed (range:1-7). The vast majority were anastomotic (10%) and “minor” non-anastomotic-biliary-strictures (NAS, 8%) requiring 1-3 stenting procedures. Four recipients (2.7%) were identified with clinically more relevant NAS, classified as diffuse necrotic or multifocal progressive with three retransplantations or relistings. Commonly used biliary parameters during NMP (i.e., bile flow, bile pH, HCO^3-, glucose) failed to identify such livers. In contrast, the mitochondrial FMN-release (start-4hrs NMP, >1000samples) into bile was the best predictor of overall and clinically impactful biliary complications as revealed by ROC-curve analysis (c-statistic: 0.82 and 0.95, Figure 1), respectively.

Conclusion:

This is the first analysis of mitochondrial biomarkers in bile during NMP. Bile FMN levels clearly identified livers with high risk to develop clinically relevant biliary strictures from those resolving with limited stenting procedures.