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There are few effective targeted strategies to reduce hepatic ischemia-reperfusion (IR) injury, a contributor to poor outcomes in liver transplantation recipients. It has been proposed that IR injury is driven by the generation of reactive oxygen species (ROS). However, recent studies implicate other mediators of the injury response, including mitochondrial metabolic dysfunction. We examined changes in global gene expression after transient hepatic ischemia and at several early reperfusion times to identify potential targets that could be used to protect against IR injury. Male Wistar rats were subjected to 30 minutes of 70% partial warm ischemia followed by 0, 0.5, 2, or 6 hours of reperfusion. RNA was extracted from the reperfused and non-ischemic lobes at each time point for microarray analysis. Identification of differentially expressed genes and pathway analysis were used to characterize IR-induced changes in the hepatic transcriptome. Changes in the reperfused lobes were specific to the various reperfusion times. We made the unexpected observation that many of these changes were also present in tissue from the paired non-ischemic lobes. However, the earliest reperfusion time, 30 minutes, showed a marked increase in the expression of a set of immediate-early genes (c-Fos, c-Jun, Atf3, Egr1) that was exclusive to the reperfused lobe. We interpreted these results as indicating that this early response represented a tissue autonomous response to reperfusion. In contrast, the changes that occurred in both the reperfused and non-ischemic lobes were interpreted as indicating a non-autonomous response resulting from hemodynamic changes and/or circulating factors. These tissue autonomous and non-autonomous responses may serve as targets to ameliorate IR injury.

Transplanting donor livers with severe macrosteatosis is associated with increased risk of primary non-function (PNF). The purpose of this study was to identify steatosis-driven biomarkers as a predisposition to severe liver damage and delayed recovery following ischemia reperfusion injury. Wistar rats were fed a methionine- and choline-deficient (MCD) diet for up to three weeks to achieve severe macrosteatosis (>90%). Animals underwent diet withdrawal to control chow and/or underwent ischemia reperfusion and partial hepatectomy injury (I/R-PHx) and reperfused out to 7 days on control chow. For animals with severe macrosteatosis, hepatic levels of IL-33 decreased while Cyclin D1 levels increased in the absence of NF-κB p65 phosphorylation. Animals with high levels of nuclear Cyclin D1 prior to I/R-PHx either did not survive or had persistent macrosteatosis after 7 days on control chow. Survival 7 days after I/R-PHx fell to 57% which correlated with increased Cyclin D1 and decreased liver IL-33 levels. In the absence of I/R-PHx, withdrawing the MCD diet normalized IL-33, Cyclin D1 levels, and I/R-PHx survival back to baseline. In transplanted grafts with macrosteatosis, higher Cyclin D1 mRNA expression was observed. Shifts in Cyclin D1 and IL-33 expression may identify severely macrosteatotic livers with increased failure risk if subjected to I/R injury. Clinical validation of the panel in donor grafts with macrosteatosis revealed increased Cyclin D1 expression corresponding to delayed graft function. This pre-surgical biomarker panel may identify the subset of livers with increased susceptibility to PNF.