Impact of Hematocrit on Normothermic Machine Perfusion of Rat Liver Grafts and Viability Testing

Abstract

Normothermic machine perfusion (NMP) has emerged as a new technique for organ storage before liver transplantation, making viability testing of the organ feasible and possibly allowing for reconditioning of marginal liver grafts. Erythrocytes form a substantial component in the composition of NMP perfusates. The optimal hematocrit (HCT) for preserving organ function during NMP and the effects of the HCT on metabolic graft function and parameters commonly used to determine organ viability are still unclear. We retrospectively analyzed our rat NMP database of 123 perfusions and identified a possible link between HCT and liver damage parameters in 34 cases following a dual perfusion protocol. To further investigate this relationship, we perfused a total of 16 rat livers for 6 h each in our rat liver NMP setup with four different HCTs at 10%, 16%, 22%, and 28%. We performed blood gas analysis and measured common viability and damage parameters such as bile production, lactate dehydrogenase (LDH), alanine aminotransferase (ALT), and aspartate aminotransferase (AST). In addition, histological analysis, as well as enzyme-linked immunosorbent assay and colorimetric assay analysis for reactive oxygen species, glycogen content, and hypoxia-inducible factor (HIF), was performed. The perfusate HCT significantly impacted ALT and LDH levels during NMP of rat liver grafts, with lower HCT values corresponding to decreased damage parameters. We investigated differentiating oxygenation levels and identified a possible cause for our observations in HIF-1α stabilization in lower HCT groups. Furthermore, we demonstrate that HCT affects key viability parameters such as lactate, pH levels, and bile production. Our findings illustrate the importance of adequate HCT levels during NMP and propose mechanisms influencing the perfusion outcome.

Impact Statement

Normothermic machine perfusion (NMP) is becoming an increasingly important tool in liver transplantation. We here show in an exploratory setting the importance of an adequate hematocrit for the outcome of liver graft perfusion and its importance in defining hepatic viability parameters during NMP.