Improvement of Normothermic Ex Vivo Machine Perfusion of Rat Liver Grafts by Dialysis and Kupffer Cell Inhibition With Glycine

Abstract

Normothermic ex vivo liver machine perfusion might be a superior preservation strategy for liver grafts from extended criteria donors. However, standardized small animal models are not available for basic research on machine perfusion of liver grafts. A laboratory‐scaled perfusion system was developed consisting of a custom‐made perfusion chamber, a pressure‐controlled roller pump, and an oxygenator. Male Wistar rat livers were perfused via the portal vein for 6 hours using oxygenated culture medium supplemented with rat erythrocytes. A separate circuit was connected via a dialysis membrane to the main circuit for plasma volume expansion. Glycine was added to the flush solution, the perfusate, and the perfusion circuit. Portal pressure and transaminase release were stable over the perfusion period. Dialysis significantly decreased the potassium concentration of the perfusate and led to significantly higher bile and total urea production. Hematoxylin‐eosin staining and immunostaining for single‐stranded DNA and activated caspase 3 showed less sinusoidal dilatation and tissue damage in livers treated with dialysis and glycine. Although Kupffer cells were preserved, tumor necrosis factor α messenger RNA levels were significantly decreased by both treatments. For proof of concept, the optimized perfusion protocol was tested with donation after circulatory death (DCD) grafts, resulting in significantly lower transaminase release into the perfusate and preserved liver architecture compared with baseline perfusion. In conclusion, our laboratory‐scaled normothermic portovenous ex vivo liver perfusion system enables rat liver preservation for 6 hours. Both dialysis and glycine treatment were shown to be synergistic for preservation of the integrity of normal and DCD liver grafts.

Dual versus single vessel normothermic ex vivo perfusion of rat liver grafts using metamizole for vasodilatation

“…The laboratory-scaled sNEVLP setup, as initially described by Gassner et al [23], consisted of a custom-made glass perfusion chamber with multiple inlets (Glass Gaßner GmbH, Munich, Germany). A flow-controlled roller pump provided a continuous flow through the portal vein.…”

“…The plasma phase was collected, and the buffy coat was withdrawn by suction. 10 mL of the erythrocyte concentrate were suspended in 35 mL of Dulbecco's Modified Eagle's Medium as used by Gassner et al [23]. 5 mL of strain specific rat plasma were added generating a total perfusion volume of 50 mL with a calculated haematocrit of 20%.…”

“…Amongst other qualities, metamizole is known to have spasmolytic effects and has been shown to effectively ameliorate arterial vasospasm of the HA of the rat [21,22]. Subsequently we compared our previously established single vessel NEVLP model (sNEVLP) to our new dNEVLP model [23].…”

Ice, ice, maybe? Is it time to ditch the igloo cooler? Benefits of machine perfusion preservation of donor hearts

“…While these systems can maintain myocardial energy stores over extended time periods, there are currently no mechanisms for clearance of waste products from the solution or replenishing the complex perfusate milieu required to maintain the organ long‐term, thus limiting perfusion of recovered donors to at most a few days with present technologies. Addition of dialysis may extend this interval further but ultimately will still be resource limited …”

“…Addition of dialysis may extend this interval further but ultimately will still be resource limited. 65 Moving from current hypothermic, static storage to truly customized and optimal donor recipient matching requires the development of organ incubators that not only meet the immediate metabolic needs but also achieve homeostasis of the organ over a period of weeks to months (Figure 2). 66 A working mode system mimicking in vivo cardiac work would be critical for maintaining donor hearts to avoid myocyte atrophy and ensure a functional organ after transplantation.…”