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Subnormothermic ex vivo liver perfusion reduces endothelial cell and bile duct injury after donation after cardiac death pig liver transplantation

Knaak JM, Spetzler VN, Goldaracena N, Boehnert MU, Bazerbachi F, Louis KS, Adeyi OA, Minkovich L, Yip PM, Keshavjee S, Levy GA, Grant DR, Selzner N,Selzner M.

Liver Transpl. 2014;20(11):1296-305.

Multi Organ Transplant Program, Department of Surgery, Toronto General Hospital, Toronto, Canada.

 

Abstract

An ischemic-type biliary stricture (ITBS) is a common feature  after liver transplantation using donation after cardiac death (DCD) grafts. We compared sequential subnormothermic ex vivo  liver  perfusion (SNEVLP; 33°C) with cold storage (CS) for the prevention of ITBS in DCD liver grafts in pig liver transplantation (n = 5 for each group). Liver grafts were stored for 10 hours at 4°C (CS) or preserved with combined 7-hour CS and 3-hour SNEVLP. Parameters of hepatocyte [aspartate aminotransferase (AST), international normalized ratio (INR), factor V, and caspase 3 immunohistochemistry], endothelial cell (EC; CD31 immunohistochemistry and hyaluronic acid), and biliary injury and function [alkaline phosphatase (ALP), total bilirubin, and bile lactate dehydrogenase (LDH)] were determined. Long-term survival (7 days) after transplantation was similar between the SNEVLP and CS groups (60% versus 40%, P = 0.13). No difference was observed between SNEVLP- and CS-treated animals with respect to the peak of serum INR, factor V, or AST levels within 24 hours. CD31 staining 8 hours after transplantation demonstrated intact EC lining in SNEVLP-treated livers (7.3 × 10(-4) ± 2.6 × 10(-4) cells/um(2)) but not in CS-treated livers (3.7 × 10(-4) ± 1.3 × 10(-4) cells/um(2) , P = 0.03). Posttransplant SNEVLP animals had decreased serum ALP and serum bilirubin levels in comparison with CS animals. In addition, LDH in bile fluid was lower in SNEVLP pigs versus CS pigs (14 ± 10 versus 60 ± 18 umol/L, P = 0.02). Bile duct histology revealed severe bile duct necrosis in 3 of 5 animals in the CS group but none in the SNEVLP group (P = 0.03). Sequential SNEVLP preservation of DCD grafts reduces bile duct and EC injuryafter liver transplantation.

© 2014 American Association for the Study of Liver Diseases.

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Significance Statement

Ischemic type biliary stricture (ITBS) is a common problem after liver transplantation with grafts obtained after cardiac death (DCD). Preventing bile duct injury in DCD liver grafts would allow us to use DCD liver transplantation more widely, and increase the donor pool for liver transplantation. Subnormothermic ex vivo liver perfusion (SNEVLP) is a novel preservation strategy for marginal grafts. During SNEVLP livers are perfused at close to physiologic temperatures with an oxygenated perfusion solution containing nutrition and mediators blocking pro-inflammatory cascades.

We compared sequential subnormothermic ex vivo liver perfusion (33°C, SNEVLP) to cold storage (CS) for the prevention of ITBS in DCD liver grafts in pig liver transplantation (n=5 for each group). Liver grafts were stored for 10hrs at 4°C (CS) or preserved using combined 7hr cold storage plus 3hr SNEVLP. Parameters of hepatocyte (AST, INR, Factor V, Caspase 3 immunohistochemistry), endothelial cell (CD31 immunohistochemistry, Hyaluronic Acid), and biliary (alkaline Phosphatase, total Bilirubin, bile LDH) injury and function were determined. Long-term survival (7days) post transplantation was similar between SNEVLP and CS groups (60% vs 40%, p=0.13). No difference was observed between SNEVLP and CS treated animals regarding peak of serum INR, Factor V, or AST levels within 24hrs. CD31 staining 8hrs post transplantation demonstrated intact endothelial cell lining in SNEVLP (7.3×10-4±2.6×10-4 cells/ um2), but not in CS treated livers (3.7×10-4±1.3×10-4 cells/um2, p=0.032). Post transplantation SNEVLP vs CS animals had decreased serum alkaline Phosphatase and serum Bilirubin levels. In addition, LDH in bile fluid was lower in SNEVLP vs CS pigs (14±10umol/L vs 60±18umol/L, p=0.02). Bile duct histology revealed severe bile duct necrosis in 3 out of 5 animals in the CS, but not in the SNEVLP group (p=0.026).

Sequential SNEVLP preservation of DCD grafts reduces bile duct and endothelial cell injury following liver transplantation. SNEVLP is an attractive preservation strategy for DCD liver grafts, with the potential to increase the DCD donor pool by minimizing ITBS after transplantation.

Figure Legend

Subnormothermic ex vivo liver perfusion circuit. The liver is perfused in a closed circuit containing an oxygenator, a centrifugal pump, a heater, and a dialysis unit.

Subnormothermic ex vivo liver perfusion reduces endothelial cell and bile duct injury after donation after cardiac death pig liver transplantation copy