Subnormothermic machine perfusion for ex vivo preservation and recovery of the human liver for transplantation.

To reduce widespread shortages, attempts are made to use more marginal livers for transplantation. Many of these grafts are discarded for fear of inferior survival rates or biliary complications. Recent advances in organ preservation have shown that ex vivo subnormothermic machine perfusion has the potential to improve preservation and recover marginal livers pretransplantation. To determine the feasibility in human livers, we assessed the effect of 3 h of oxygenated subnormothermic machine perfusion (21°C) on seven livers discarded for transplantation. Biochemical and microscopic assessment revealed minimal injury sustained during perfusion. Improved oxygen uptake (1.30 [1.11-1.94] to 6.74 [4.15-8.16] mL O2 /min kg liver), lactate levels (4.04 [3.70-5.99] to 2.29 [1.20-3.43] mmol/L) and adenosine triphosphate content (45.0 [70.6-87.5] pmol/mg preperfusion to 167.5 [151.5-237.2] pmol/mg after perfusion) were observed. Liver function, reflected by urea, albumin and bile production, was seen during perfusion. Bile production increased and the composition of bile (bile salts/phospholipid ratio, pH and bicarbonate concentration) became more favorable. In conclusion, ex vivo subnormothermic machine perfusion effectively maintains liver function with minimal injury and sustains or improves various hepatobiliary parameters postischemia.

Perfusion defatting at subnormothermic temperatures in steatotic rat livers.

Hepatic steatosis is a major risk factor in liver transplantation. Use of machine perfusion to reduce steatosis has been reported previously at normothermic (37°C) temperatures, with minimal media as well as specialized defatting cocktails. In this work, we tested if subnormothermic (room temperature) machine perfusion, a more practical version of machine perfusion approach that does not require temperature control or oxygen carriers, could also be used to reduce fat content in steatotic livers. Steatotic livers recovered from obese Zucker rats were perfused for 6 hours. A significant increase of very low density lipoprotein (VLDL) and triglyceride (TG) content in perfusate, with or without a defatting cocktail, was observed although the changes in histology were minimal and changes in intracellular TG content were not statistically significant. The oxygen uptake rate, VLDL secretion, TG secretion, and venous resistance were similar in both groups. This study confirms lipid export during subnormothermic machine perfusion; however, the duration of perfusion necessary appears much higher than required in normothermic perfusion.

Hepatocyte viability and adenosine triphosphate content decrease linearly over time during conventional cold storage of rat liver grafts.

INTRODUCTION: The gold standard in organ preservation is static cold storage (SCS) using University of Wisconsin solution (UW). Although it is well-known that there is a finite limit to SCS preservation, and that there is a correlation between the adenosine triphosphate (ATP) levels and organ function post-preservation, a quantitative relationship has not been established, which is important in understanding the fundamental limitations to preservation, minimizing cold ischemic injury, and hence maximizing use of the donor organ pool. AIM: This study determines the time limits of cellular viability and metabolic function during SCS, and characterizes the relationship between cellular viability and energetic state using clinically relevant techniques in organ preservation. METHODS: Rat livers were procured and stored using conventional storage in UW solution at 4 °C. Viability was assessed by determining the amount of viable hepatocytes and intracellular ATP content after 0, 24, 48, 72, and 120 hours of storage. RESULTS: Numbers of viable hepatocytes that were isolated from these livers decreased steadily during SCS. After 5 days, viable hepatocytes decreased from 25.95 × 10(6) to 0.87 × 10(6) cells/gram tissue. Intracellular ATP content decreased from 9.63 to 0.93 moles/g tissue. Statistical analysis of variance established a linear relation for both parameters as a function of time (P < .05). CONCLUSION: The linear correlation between hepatocyte viability, ATP content, and storage time suggests a shared physiological foundation. These findings confirm ATP as direct predictor for organ quality in the context of liver preservation, which will aid quantitative assessment of donor organs for various applications.

Diluted blood reperfusion as a model for transplantation of ischemic rat livers: alanine aminotransferase is a direct indicator of viability.

Donors after cardiac death present a significant pool of untapped organs for transplantation, and use of machine perfusion strategies has been an active focus area in experimental transplantation. However, despite 2 decades of research, a gold standard has yet to emerge for machine perfusion systems and protocols. Whole blood reperfusion has been used as a surrogate for organ transplantation, especially as a model for the short-term response posttransplantation, and for optimization of perfusion systems. Although it is known that there is a strong correlation between liver function in whole-blood reperfusion and survival, the exact nature of these correlations, and to what extent they can be considered as an indicator of viability for transplantation/recipient survival, remain unclear. In this work, we demonstrate that diluted whole-blood reperfusion can be used as a direct model for transplantation of ischemic rat liver grafts. Specifically, we show that recipient survival can be predicted based simply on the value of alanine aminotransferase during perfusion, providing quantitative criteria of viability for use in this animal model. These results indicate that in the rat model graft survival is highly correlated with hepatocellular damage.

Sequential cold storage and normothermic perfusion of the ischemic rat liver.

Extending transplant criteria to include livers obtained from donors after cardiac death (DCD) could increase the liver donor pool, but conventional simple cold storage of these ischemic organs can lead to poor graft function after transplantation. Experimental normothermic machine perfusion has previously proven to be useful for the recovery and preservation of DCD livers, but it is more complicated than conventional cold storage, and, therefore, is perhaps not practical during the entire preservation period. In clinical situations, the combined use of simple cold storage and normothermic perfusion preservation of DCD livers might be more realistic, but even a brief period of cold storage prior to normothermic preservation has been suggested to have a negative impact on graft viability. In this study we show that rat livers subjected to 45 minutes of ex vivo warm ischemia followed by 2 hours of simple cold storage can be reclaimed by 4 hours of normothermic machine perfusion. These livers could be orthotopically transplanted into syngeneic recipients with 100% survival after 4 weeks (N = 10), similar to the survival of animals that received fresh livers that were stored on ice in University of Wisconsin (UW) solution for 6 hours (N = 6). On the other hand, rats that received ischemic livers preserved on ice in UW solution for 6 hours (N = 6) all died within 12 hours after transplantation. These results suggest that normothermic perfusion can be used to reclaim DCD livers subjected to an additional period of cold ischemia during hypothermic storage.