Manipulation of Lipid Metabolism During Normothermic Machine Perfusion: Effect of Defatting Therapies on Donor Liver Functional Recovery.

Yuri L Boteon,Darius F Mirza,Simon C Afford,Gary Reynolds,Ricky H Bhogal,Amanda P C S Boteon,Hynek Mergental,Joseph Attard,Lorraine Wallace,Stefan Hubscher

doi:10.1002/lt.25439

Abstract

Strategies to increase the use of steatotic donor livers are required to tackle the mortality on the transplant waiting list. We aimed to test the efficacy of pharmacological enhancement of the lipid metabolism of human livers during ex situ normothermic machine perfusion to promote defatting and improve the functional recovery of the organs. Because of steatosis, 10 livers were discarded and were allocated either to a defatting group that had the perfusate supplemented with a combination of drugs to enhance lipid metabolism or to a control group that received perfusion fluid with vehicle only. Steatosis was assessed using tissue homogenate and histological analyses. Markers for lipid oxidation and solubilization, oxidative injury, inflammation, and biliary function were evaluated by enzyme‐linked immunosorbent assay, immunohistochemistry, and in‐gel protein detection. Treatment reduced tissue triglycerides by 38% and macrovesicular steatosis by 40% over 6 hours. This effect was driven by increased solubility of the triglycerides (P = 0.04), and mitochondrial oxidation as assessed by increased ketogenesis (P = 0.008) and adenosine triphosphate synthesis (P = 0.01) were associated with increased levels of the enzymes acyl‐coenzyme A oxidase 1, carnitine palmitoyltransferase 1A, and acetyl‐coenzyme A synthetase. Concomitantly, defatted livers exhibited enhanced metabolic functional parameters such as urea production (P = 0.03), lower vascular resistance, lower release of alanine aminotransferase (P = 0.049), and higher bile production (P = 0.008) with a higher bile pH (P = 0.03). The treatment down‐regulated the expression of markers for oxidative injury as well as activation of immune cells (CD14; CD11b) and reduced the release of inflammatory cytokines in the perfusate (tumor necrosis factor α; interleukin 1β). In conclusion, pharmacological enhancement of intracellular lipid metabolism during normothermic machine perfusion decreased the lipid content of human livers within 6 hours. It also improved the intracellular metabolic support to the organs, leading to successful functional recovery and decreased expression of markers of reperfusion injury.

Highlights

Manipulation of Lipid Metabolism During Normothermic Machine Perfusion: Effect of Defatting Therapies on Donor Liver Functional Recovery
Steatosis is caused by the abnormal metabolism of fatty acids (FAs) in hepatocytes, resulting in Abbreviations: 4-HNE, 4-hydroxynonenal; 8-HOdG, 8-hydroxy-2deoxyguanosine; ACOX1, acyl-coenzyme A oxidase 1; ALT, alanine aminotransferase; AST, aspartate aminotransferase; ATP, adenosine triphosphate; AUC, area under the curve; cAMP, cyclic adenosine monophosphate; CAR, constitutive androstane receptor; cold ischemia time (CIT), cold intracytoplasmic accumulation of triacylglycerol as lipid droplets (LDs).(1) In the context of organ donation, livers with large intracytoplasmic LDs displacing the cell nucleus, ie, macrovesicular steatosis (MaS), are more vulnerable to ischemia/reperfusion injury (IRI) during standard static cold storage (SCS)
Steatosis has become the leading reason for surgeons to decline donor livers for transplantation, which, in turn, is worsening the growing discrepancy between organ availability and the increasing number of patients on transplant waiting lists.[14,15] We have shown that the delivery of a pharmacological intervention during normothermic machine perfusion (NMP) was able to decrease the fat content of whole human livers within 6 hours

Summary

Introduction

Manipulation of Lipid Metabolism During Normothermic Machine Perfusion: Effect of Defatting Therapies on Donor Liver Functional Recovery. Steatosis is caused by the abnormal metabolism of fatty acids (FAs) in hepatocytes, resulting in Abbreviations: 4-HNE, 4-hydroxynonenal; 8-HOdG, 8-hydroxy-2deoxyguanosine; ACOX1, acyl-coenzyme A oxidase 1; ALT, alanine aminotransferase; AST, aspartate aminotransferase; ATP, adenosine triphosphate; AUC, area under the curve; cAMP, cyclic adenosine monophosphate; CAR, constitutive androstane receptor; CIT, cold intracytoplasmic accumulation of triacylglycerol as lipid droplets (LDs).(1) In the context of organ donation, livers with large intracytoplasmic LDs displacing the cell nucleus, ie, macrovesicular steatosis (MaS), are more vulnerable to ischemia/reperfusion injury (IRI) during standard static cold storage (SCS) This is predominantly due to impaired mitochondrial function, poor microcirculation, and exaggerated inflammatory response leading to tissue damage.[2] The exacerbated IRI is associated with impaired early functional recovery and a high risk of early allograft. Ischemia time; CoA, coenzyme A; CPT1A, carnitine palmitoyltransferase form 1A; DBD, donation after brain death; DCD, donation after circulatory death; DMSO, dimethyl sulfoxide; ECD, extended criteria donor; ET, Eurotransplant; FA, fatty acid; GGT, gamma-glutamyl transpeptidase; H & E, hematoxylin-eosin; HA, hepatic artery; HDL, high-density lipoprotein; IL, interleukin; IQR, interquartile range; IRI, ischemia/reperfusion injury; IRS, modified immunoreactive score; LD, lipid droplet; MaS, macrovesicular steatosis; MiS, microvesicular steatosis; NHSBT, National Health Service Blood and Transplant; NMP, normothermic machine perfusion; PAS, periodic acid–Schiff; PCO2, partial pressure of carbon dioxide; PPAR, peroxisome proliferatoractivated receptor; P-TG, perfusate triglyceride; PXR, pregnane X receptor; PV, portal vein; ROS, reactive oxygen species; SCS, static cold storage; TG, triglyceride; TKB, total ketone bodies; TNF-α, tumor necrosis factor α; T-TG, tissue triglyceride; VLDL, very low density lipoprotein; WIT, warm ischemia time

Objectives

Methods

Results

Conclusion